Benzimidazolone Compounds Having 5-HT4 Receptor Agonistic Activity

ABSTRACT

This invention provides a compound of the formula (I): 
     
       
         
         
             
             
         
       
     
     or a pharmaceutically acceptable salt thereof, and compositions containing such compounds and the use of such compounds for the manufacture of medicament for gastroesophageal reflux disease, gastrointestinal disease, gastric motility disorder, non-ulcer dyspepsia, functional dyspepsia, irritable bowel syndrome (IBS), constipation, dyspepsia, esophagitis, gastroesophageral disease, nausea, central nervous system disease, Alzheimer&#39;s disease, cognitive disorder, emesis, migraine, neurological disease, pain, cardiovascular disorders, cardiac failure, heart arrhythmia, diabetes and apnea syndrome. 
     These compounds have 5-HT 4  receptor agonistic activity, and thus are useful for the treatment of gastroesophageal reflux disease, non-ulcer dyspepsia, functional dyspepsia, irritable bowel syndrome or the like in mammalian, especially humans.

TECHNICAL FIELD

This invention relates to novel benzimidazolone compounds. Thesecompounds have selective 5-HT₄ receptor agonistic activity. The presentinvention also relates to a pharmaceutical composition, a method oftreatment and use comprising the above compounds for the treatment ofdisease conditions mediated by 5-HT₄ receptor activity.

BACKGROUND ART

In general, 5-HT₄ receptor agonists are found to be useful for thetreatment of a variety of diseases such as gastroesophageal refluxdisease, gastrointestinal disease, gastric motility disorder, non-ulcerdyspepsia, functional dyspepsia, irritable bowel syndrome (IBS),constipation, dyspepsia, esophagitis, gastroesophageral disease, nausea,central nervous system disease, Alzheimer's disease, cognitive disorder,emesis, migraine, neurological disease, pain, cardiovascular disorders,cardiac failure, heart arrhythmia, diabetes and apnea syndrome (SeeTiPs, 1992, 13, 141; Ford A. P. D. W. et al., Med. Res. Rev., 1993, 13,633; Gullikson G. W. et al., Drug Dev. Res., 1992, 26, 405; Richard M.Eglen et al, TiPS, 1995, 16, 391; Bockaert J. Et al., CNS Drugs, 1, 6;Romanelli M. N. et al., Arzheim Forsch./Drug Res., 1993, 43, 913;Kaumann A. et al., Naunyn-Schmiedeberg's. 1991, 344, 150; and RomanelliM. N. et al., Arzheim Forsch./Drug Res., 1993, 43, 913). Also, Mosaprideis known to be useful for the treatment of diabetes.

It would be desirable if there were provided 5-HT₄ receptor agonistswhich have more 5HT₄ receptor agonistic activities.

U.S. Pat. No. 5,223,511 discloses benzimidazole compounds as 5-HT₄receptor antagonists. Especially, compounds represented by the followingformula is disclosed:

WO93/18027 discloses benzimidazolone compounds as 5-HT₄ receptorantagonists. Especially, compounds represented by the following formulais disclosed:

WO99/17772 discloses benzimidazolone compounds as 5-HT₄ receptoragonists and/or antagonists. Especially, compounds represented by thefollowing formula is disclosed:

WO94/00449 discloses benzimidazolone compounds as 5-HT₄ agonists orantagonists and/or 5-HT₃ antagonists. Especially, compounds representedby the following formula is disclosed:

There is a need to provide new 5-HT₄ agonists that are good drugcandidates. In particular, preferred compounds should bind potently tothe 5-HT₄ receptor whilst showing little affinity for other receptorsand show functional activity as agonists. They should be well absorbedfrom the gastrointestinal tract, be metabolically stable and possessfavorable pharmacokinetic properties. When targeted against receptors inthe central nervous system they should cross the blood brain barrierfreely and when targeted selectively against receptors in the peripheralnervous system they should not cross the blood brain barrier. Theyshould be non-toxic and demonstrate few side-effects. Furthermore, theideal drug candidate will exist in a physical form that is stable,non-hygroscopic and easily formulated.

BRIEF DISCLOSURE OF THE INVENTION

It has now surprisingly been found that compounds of this invention havestrong selective 5-HT₄ agonistic activity, and thus are useful for thetreatment of disease conditions mediated by 5-HT₄ activity such asgastroesophageal reflux disease, gastrointestinal disease, gastricmotility disorder, non-ulcer dyspepsia, functional dyspepsia, irritablebowel syndrome (IBS), constipation, dyspepsia, esophagitis,gastroesophageral disease, nausea, central nervous system disease,Alzheimer's disease, cognitive disorder, emesis, migraine, neurologicaldisease, pain, cardiovascular disorders, cardiac failure, heartarrhythmia, diabetes and apnea syndrome (especially caused by an opioidadministration).

Further, the compounds of the present invention show a reduced QTprolongation by introducing a polar group into R³ of the formula (I). QTprolongation is known to have a potential liability to produce fatalcardiac arrhythmias of Torsades de Pointes (TdP). The ability to prolongthe cardiac action potential duration was identified as being due to anaction at the HERG potassium channel. For example, drugs withdrawn fromthe market due to QT prolongation, such as Cisapride and Terfenadine,are known to be potent HERG potassium channel blocker (Expert Opinion ofPharmacotherapy.; 2, pp 947-973, 2000) Inhibitory activity at HERGchannel was estimated from affinity for HERG type potassium channel wasinvestigated by checking [³H]dofetilide binding, which can predictinhibitory activity at HERO channel (Eur. J. Pharmacol., 430, pp147-148, 2001).

The compounds of the present invention may show less toxicity, goodabsorption, distribution, good solubility, low protein binding affinity,less drug-drug interaction, and good metabolic stability.

The present invention provides a compound of the following formula (1)or a pharmaceutically acceptable salt thereof.

wherein

-   Het represents a heterocyclic group having one nitrogen atom, to    which B binds directly, and from 4 to 7 carbon atoms, and said    heterocyclic group being unsubstituted or substituted by 1 to 4    substituents independently selected from the group consisting of    substituents α¹;-   A represents an alkylene group having from 1 to 4 carbon atoms;-   B represents a covalent bond or an alkylene group having from 1 to 5    carbon atoms, and said alkylene group being unsubstituted or    substituted by an oxo group when R³ represents a heterocyclic group;-   R¹ represents an isopropyl group or a cyclopentyl group;-   R² independently represents a halogen atom or an alkyl group having    from 1 to 4 carbon atoms; m is 0, 1, 2, 3 or 4; and-   R³ represents    -   (i) a cycloalkyl group having from 3 to 8 carbon atoms, and said        cycloalkyl group being substituted by 1 to 5 substituents        independently selected from the group consisting of substituents        α², or    -   (ii) a heterocyclic group having from 3 to 8 atoms, and said        heterocyclic group being unsubstituted or substituted by 1 to 5        substituents independently selected from the group consisting of        substituents β,-   said substituents α¹ are independently selected from a hydroxy group    and an amino group;-   said substituents α² are independently selected from a hydroxy    group, an amino group, a hydroxy-substituted alkyl group having from    1 to 4 carbon atoms, a carboxyl group and an alkoxy group having    from 1 to 4 carbon atoms; and-   said substituents β are independently selected from a hydroxy group,    a hydroxy-substituted alkyl group having from 1 to 4 carbon atoms, a    carboxyl group, an amino group, an alkyl group having from 1 to 4    carbon atoms, an amino-substituted alkyl group having from 1 to 4    carbon atoms and a carbamoyl group.

The invention also provides a compound of the formula (I):

or a pharmaceutically acceptable salt thereof, wherein:

-   Het represents a heterocyclic group having one nitrogen atom, to    which B binds directly, and from 4 to 7 carbon atoms, and said    heterocyclic group being unsubstituted or substituted by 1 to 4    substituents independently selected from the group consisting of    substituents α¹:-   A represents an alkylene group having from 1 to 4 carbon atoms;-   B represents a covalent bond or an alkylene group having from 1 to 5    carbon atoms, and said alkylene group being unsubstituted or    substituted by an oxo group when R³ represents a heterocyclic group;-   R¹ represents an isopropyl group or a cyclopentyl group;-   R² independently represents a halogen atom or an alkyl group having    from 1 to 4 carbon atoms; m is 0, 1, 2, 3 or 4; and-   R³ represents    -   (i) a cycloalkyl group having from 3 to 8 carbon atoms, and said        cycloalkyl group being substituted by 1 to 5 substituents        independently selected from the group consisting of substituents        α², or    -   (ii) a heterocyclic group having from 3 to 8 atoms, and said        heterocyclic group being unsubstituted or substituted by 1 to 5        substituents independently selected from the group consisting of        substituents β,-   said substituents α¹ are independently selected from a hydroxy group    and an amino group;-   said substituents α² are independently selected from a hydroxy    group, an amino group, a hydroxy-substituted alkyl group having from    1 to 4 carbon atoms, a carboxyl-substituted alkyl group having 1 to    4 carbon atoms, a carboxyl group and an alkoxy group having from 1    to 4 carbon atoms; and-   said substituents β are independently selected from a hydroxy group,    a hydroxy-substituted alkyl group having from 1 to 4 carbon atoms, a    carboxyl-substituted alkyl group having 1 to 4 carbon atoms, a    carboxyl group, an amino group, an alkyl group having from 1 to 4    carbon atoms, an amino-substituted alkyl group having from 1 to 4    carbon atoms and a carbamoyl group.

Also, the present invention provides a pharmaceutical composition forthe treatment of disease conditions mediated by 5-HT₄ receptor, in amammalian subject, which comprises administering to said subject atherapeutically effective amount of a compound of formula (I) orpharmaceutically acceptable salts thereof.

Further, the present invention also provides a pharmaceuticalcomposition for the treatment of diseases selected from gastroesophagealreflux disease, gastrointestinal disease, gastric motility disorder,non-ulcer dyspepsia, functional dyspepsia, irritable bowel syndrome(IBS), constipation, dyspepsia, esophagitis, gastroesophageral disease,nausea, central nervous system disease, Alzheimer's disease, cognitivedisorder, emesis, migraine, neurological disease, pain, cardiovasculardisorders, cardiac failure, heart arrhythmia, diabetes and apneasyndrome, or the like, which comprises a therapeutically effectiveamount of the benzimidazolone compound of formula (I) or itspharmaceutically acceptable salt together with a pharmaceuticallyacceptable carrier.

Also, the present invention provides a method of the treatment of amammal, including a human, to treat a disease conditions mediated by5-HT₄ receptor, in a mammalian subject, which comprises administering tosaid subject a therapeutically effective amount of a compound of formula(I) or pharmaceutically acceptable salts thereof. Further, the presentinvention provides a method for the treatment of the disease conditionsas mentioned above. Furthermore, the present invention provides use ofthe compound of formula (I) or pharmaceutically acceptable salts thereofin the manufacture of a medicament for the treatment of diseaseconditions mediated by 5-HT₄ receptor activity, in a mammalian subject.The conditions mediated by 5-HT₄ receptor activity include thosediseases or disorders described as above.

Also, the present invention provides a compound of the following formula(2-A′) or a salt thereof:

wherein

-   R^(a) represents a hydrogen atom or a N-protecting group;-   Het represents a heterocyclic group having one nitrogen atom, to    which B binds directly, and from 4 to 7 carbon atoms, and said    heterocyclic group being unsubstituted or substituted by 1 to 4    substituents independently selected from the group consisting of    substituents α¹;-   A represents an alkylene group having from 1 to 4 carbon atoms;-   B represents a covalent bond or an alkylene group having from 1 to 5    carbon atoms, and said alkylene group being unsubstituted or    substituted by an oxo group when R³ represents a heterocyclic group;-   R³ represents    -   (i) a cycloalkyl group having from 3 to 8 carbon atoms, and said        cycloalkyl group being substituted by 1 to 5 substituents        independently selected from the group consisting of substituents        α², or    -   (ii) a heterocyclic group having from 3 to 8 atoms, and said        heterocyclic group being unsubstituted or substituted by 1 to 5        substituents independently selected from the group consisting of        substituents β,-   said substituents α¹ are independently selected from a hydroxy group    and an amino group;-   said substituents α² are independently selected from a hydroxy    group, an amino group, a hydroxy-substituted alkyl group having from    1 to 4 carbon atoms, a carboxyl group and an alkoxy group having    from 1 to 4 carbon atoms; and-   said substituents β are independently selected from a hydroxy group,    a hydroxy-substituted alkyl group having from 1 to 4 carbon atoms, a    carboxyl group, an amino group, an alkyl group having from 1 to 4    carbon atoms, an amino-substituted alkyl group having from 1 to 4    carbon atoms and a carbamoyl group,

DETAILED DESCRIPTION OF THE INVENTION

As used herein, the term “heterocyclic” of “Het” means a heterocyclicgroup having one nitrogen atom and from 4 to 7 carbon atoms such as

As used herein, the term “alkylene” in “A” means straight or branchedchain saturated radicals having 1 to 4 carbon atoms, including, but notlimited to methylene, ethylene, n-propylene, isopropylene, n-butylene,isobutylene, sec-butylene, cert-butylene. The “alkylene” in “A”represents preferably a methylene group, an ethylene group or apropylene group; more preferably a methylene group or an ethylene group;most preferably a methylene group.

As used herein, the term “alkylene” in “B” means straight or branchedchain saturated radicals having 1 to 5 carbon atoms, including, but notlimited to methylene, ethylene, n-propylene, isopropylene, n-butylene,isobutylene, sec-butylene, tert-butylene, n-pentylene, isopentylene,sec-pentylene, tert-pentylene. The “alkylene” in “B” representspreferably an alkylene group having from 1 to 4 carbon atoms; morepreferably an alkylene group having from 1 to 3 carbon atoms; much morepreferably a methylene group or an ethylene group; further morepreferably a methylene group.

As used herein, the term “halogen” in “R²” means fluoro, chloro, bromoand iodo, preferably fluoro or chloro.

As used herein, the term “alkyl” in “R²”; “alkyl” of “ahydroxy-substituted alkyl group” and “an alkoxy group having from 1 to 4carbon atoms” in “substituents α²”; “alkyl” in “substituents β”; and“alkyl” of “a hydroxy-substituted alkyl group” and “an amino-substitutedalkyl group” in “substituents β” mean straight or branched chainsaturated radicals having 1 to 4 carbon atoms, including, but notlimited to methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl,sec-butyl, tert-butyl.

As used herein, the term “cycloalkyl” in “R³” means cyclic alkyl grouphaving 3 to 8 carbon atoms, such as cyclopropyl, cyclobutyl,cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, and etc.

As used herein, the term “heterocyclic” of “R³” means a heterocyclicring which has one or more hetero atoms in the ring, preferably has 2 to6 carbon atoms and 1 to 3 heteroatoms, including aziridinyl, azetidinyl,piperidinyl, morpholinyl (including morpholino), pyrrolidinyl,pyrazolidinyl, piperazinyl, tetrahydropyrazolyl, pyrazolinyl,tetrahydropyranyl and etc.

The term “treating”, as used herein, refers to reversing, alleviating,inhibiting the progress of, or preventing the disorder or condition towhich such term applies, or one or more symptoms of such disorder orcondition. The term “treatment” as used herein refers to the act oftreating, as “treating” is defined immediately above.

A preferred compound of formula (1) of this invention is that whereinHet represents a heterocyclic group selected from

said heterocyclic group being unsubstituted or substituted by 1 to 3substituents independently selected from the group consisting ofsubstituents α¹; and

-   A represents an alkylene group having from 1 to 3 carbon atoms.

A more preferred compound of formula (I) of this invention is thatwherein

-   Het represents a group of formula

and this group being unsubstituted or substituted by one substituentselected from the group consisting of substituents α¹;

-   A represents an alkylene group having from 1 to 2 carbon atoms;-   B represents an alkylene group having from 1 to 4 carbon atoms, and    said alkylene group being unsubstituted or substituted by an oxo    group when R³ represents a heterocyclic group;-   R² independently represents a halogen atom or an alkyl group having    from 1 to 2 carbon atoms; m is 0, 1 or 2; and-   R³ represents    -   (i) a cycloalkyl group having from 4 to 7 carbon atoms, and said        cycloalkyl group being substituted by 1 to 3 substituents        independently selected from the group consisting of substituents        α², or    -   (ii) a heterocyclic group having from 4 to 7 atoms, and said        heterocyclic group being unsubstituted or substituted by 1 to 3        substituents independently selected from the group consisting of        substituents β.

Also, a further preferred compound of formula (I) of this invention isthe compound or its pharmaceutically acceptable salt wherein

-   Het represents a group of formula

and this group being unsubstituted or substituted by one substituentselected from the group consisting of substituents α¹;

-   A represents a methylene group;-   B represents an alkylene group having from 1 to 2 carbon atoms;-   R¹ represents an isopropyl group;-   R² independently represents a fluorine atom, a chlorine atom or a    methyl; and-   R³ represents    -   (i) a cycloalkyl group having from 5 to 7 carbon atoms, and said        cycloalkyl group being substituted by 1 to 2 substituents        independently selected from the group consisting of substituents        α², or    -   (ii) a heterocyclic group having from 5 to 7 atoms, and said        heterocyclic group being unsubstituted or substituted by 1 to 2        substituents independently selected from the group consisting of        substituents β,-   said substituents α² are independently selected from a hydroxy    group, an amino group and an alkoxy group having from 1 to 2 carbon    atoms; and-   said substituents β are independently selected from a hydroxy group,    a hydroxy-substituted alkyl group having from 1 to 2 carbon atoms, a    carboxyl group, an amino group, an amino-substituted alkyl group    having from 1 to 2 carbon atoms and a carbamoyl group.

A further preferred compound of formula (I) of this invention is thecompound or its pharmaceutically acceptable salt wherein

-   Het represents a group of formula

-   A represents a methylene group;-   B represents a methylene group;-   R¹ represents an isopropyl group;-   R² represents a fluorine atom; m is 0 or 1; and-   R³ represents    -   (i) a cycloalkyl group having from 5 to 6 carbon atoms, and said        cycloalkyl group being substituted by 1 to 2 substituents        independently selected from the group consisting of substituents        α², or    -   (ii) a heterocyclic group having from 5 to 6 atoms, and said        heterocyclic group being unsubstituted or substituted by 1 to 2        substituents independently selected from the group consisting of        substituents β,-   said substituents α² are independently selected from a hydroxy group    and an amino group; and-   said substituents β are independently selected from a hydroxy group    and an amino group.

A further preferred compound of formula (I) of this invention is thecompound or its pharmaceutically acceptable salt, wherein

-   Het represents a group of formula

-   A represents a methylene group;-   B represents a methylene group;-   R¹ represents an isopropyl group;-   R² represents a fluorine atom; m is 0; and-   R³ represents    -   (i) a cyclohexyl group substituted by 1 to 2 substituents        independently selected from a hydroxy group or an amino group,        or    -   (ii) a heterocyclic group having from 6 atoms, and said        heterocyclic group being substituted by a hydroxy group or an        amino group.

Most preferred compounds of formula (I) of this invention is thecompound or its pharmaceutically acceptable salt, wherein

-   Het represents a group of formula

-   A represents a methylene group;-   B represents a methylene group;-   R¹ represents an isopropyl group;-   R² represents a fluorine atom; m is 0; and-   R³ represents    -   (i) a cyclohexyl group substituted by 1 or 2 hydroxy group        (especially dihydroxycyclohexyl), or    -   (ii) a tetrahydropyran group substituted by 1 or 2 hydroxy group        (especially hydroxytetrahydropyranyl).

In the compounds of formula (I) or the pharmaceutically acceptable salt,R² preferably represents a fluorine atom, a chlorine atom, a methylgroup or an ethylene group; more preferably a fluorine atom, a chlorineatom, a methyl group; most preferably a fluorine atom.

In the compounds of formula (I) or the pharmaceutically acceptable salt,m is preferably 0, 1 or 2; more preferably 0 or 1; much more preferably0.

Preferred individual compound of this invention is:

-   N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide;-   N-({1-[(trans-1,4-dihydroxyhexyl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide;-   N-({1-[(cis-1,4-dihydroxyhexyl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide;-   6-fluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide;-   4-{[4-({[(3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazol-1-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-carboxylic    acid;-   1-{[4-({[(3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazol-1-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl}cyclopentanecarboxylic    acid; and-   1-{[4-({[(3-isopropyl-2-oxo-2,3-dihydro-1H    -benzimidazol-1-yl)carbonyl]amino}methyl)piperidin-1-yl]methyl    cyclobutanecarboxylic acid;    or a pharmaceutically acceptable salt thereof.

A preferred compound of formula (2-A′) of this invention is that wherein

-   R^(a) represents a hydrogen atom or a t-butoxycarbonyl group;-   Het represents a group of formula

-   A represents an methylene group; B represents an methylene group;    and-   R³ represents hydroxytetrahydropyranyl or dihydroxycyclohexyl.

General Synthesis

The compounds of the present invention may be prepared by a variety ofprocesses well known for the preparation of compounds of this type, forexample as shown in the following reaction Schemes. Unless otherwiseindicated R¹ through R³ and m in the following reaction Schemes anddiscussion are defined as above. The term “protecting group”, as usedhereinafter, means a hydroxy or amino protecting group which is selectedfrom typical hydroxy or amino protecting groups described in ProtectiveGroups in Organic Synthesis edited by T. W. Greene et al. (John Wiley &Sons, 1991); All starting materials in the following general synthesesmay be commercially available or obtained by conventional methods knownto those skilled in the art.

The compound of formula (I), wherein Het

is prepared by the following synthesis. And the compound of formula (I),wherein Het is other than

can be prepared by a similar manner or a method known to a skilledperson.

In Steps 1a, 1b, 1d, 2a, 2c, 2e, 3a, 3c, 3d of the following schemes,each reaction is preferably carried out in the presence of a base. Thereis no particular restriction on the nature of the bases used, and anybase commonly used in reactions of this type may equally be used here.The base employed includes, for example, alkali metal hydroxides, suchas lithium hydroxide, sodium hydroxide and potassium hydroxide; alkalimetal carbonates such as sodium carbonate and potassium carbonate;alkali metal hydrides such as sodium hydride, potassium hydride andlithium hydride; alkali metal alkoxides such as sodium methoxide, sodiumethoxide, potassium t-butoxide and lithium methoxide; alkyllithiums suchas butyllithium and methyllithium; lithium amides such as lithiumdiethylamides, lithium diisopropylamide and lithiumbis(trimethylsilyl)amide; alkali metal hydrogencarbonates such as sodiumhydrogencarbonate and potassium hydrogencarbonate; and tertiary organicamines such as triethylamine, dimethylaniline, pyridine,4-dimethylaminopyridine, 1,5-diazabicyclo[4.3.0]non-5-ene,1,8-diazabicyclo[5.4.0]undec-7-ene and N,N-diisopropylethylamine.

Synthesis of Benzimidazolone (1-A):

The following reaction Schemes illustrate the preparation ofbenzimidazolone compounds of formula 1-A.

In the above formulae, Z represents ‘halo’, such as a chlorine, bromineor iodine atom.

Step 1a

In step 1a, an amine compound of formula 1-3 can be prepared by thereductive amination of the alkanone compound (having from 1 to 4 carbonatoms) with an amine compound of formula 1-1 in the presence or absenceof a reducing agent or a metal agent in an inert solvent.

The reaction is normally and preferably effected in the presence of asolvent. There is no particular restriction on the nature of the solventto be employed, provided that it has no adverse effect on the reactionor on the reagents involved and that it can dissolve the reagents, atleast to some extent. Examples of suitable aqueous or non-aqueousorganic solvents include: alcohols, such as methanol, ethanol orisopropanol; ethers, such as tetrahydrofuran(THF), dimethoxyethane ordioxane; acetonitrile; N,N′-dimethylformamide; dimethylsulfoxide; aceticacid; and halogenated hydrocarbon, such as dichloromethane,dichloroethane or chloroform.

The reaction can take place over a wide range of temperatures, and theprecise reaction temperature is not critical to the invention. Thepreferred reaction temperature will depend upon such factors as thenature of the solvent, and the starting material or reagent used.However, in general, it is convenient to carry out the reaction withreducing agents at a temperature of from −78° C. to 100° C., morepreferably from about −20° C. to 60° C. The time required for thereaction may also vary widely, depending on many factors, notably thereaction temperature and the nature of the reagents and solventemployed. However, provided that the reaction is effected under thepreferred conditions outlined above, a period of 5 minutes to 1 week,more preferably 30 minutes to 24 hours, will usually suffice. In thecase of the reaction with metal reagents, it is convenient to carry outthe reaction at a temperature of from 20° C. to 100° C., preferably fromabout 20° C. to 60° C. for 10 minutes to 48 hours, preferably 30 minutesto 24 hours.

Suitable reducing reagents are those typically used in the reductionincluding, for example, sodium borohydride, sodium cyanoborohydride orsodium triacetoxyborohydride.

The combination of metal reagents and hydrogen gas can be also employedas reducing reagent. Example of suitable metal reagents includepalladium-carbon, palladiumhydroxide-carbon, platinumoxide,platinum-carbon, ruthenium-carbon, rhodium-aluminumoxide andtris[triphenyphosphine]rhodiumchloride. The reduction with metalreagents may be carried out under hydrogen atmosphere at a pressureranging from 1 to 100 atm, preferably from 1 to 10 atm.

This reduction can be carried out after formation of the correspondingenamine of the alkanone compound or imine of the alkanone compound in areaction-inert solvent such as benzene, toluene, or xylene at atemperature in the range from 20 to 130° C. for 1 hour to 1 week.

Alternatively, the compound of formula 1-3 can be prepared by alkylationof the compound of formula 1-1 with an alkyl halide of formula of Z—R¹wherein Z is halo (halo is chloro, bromo, or iodo) as essentially thesame condition as below (Step 1d), preferably in the presence of a base.

Step 1b

In this step, a compound of formula 1-3 can be prepared by alkylation ofa compound of formula 1-2 with compound of formula R¹—NH₂.

The reaction can take place over a wide range of temperatures, and theprecise reaction temperature is not critical to the invention. Thepreferred reaction temperature will depend upon such factors as thenature of the solvent, and the starting material or reagent used.However, in general, it is convenient to carry out the reaction at atemperature of from 0° C. to 150° C., more preferably from 20° C. to120° C. The time required for the reaction may also vary widely,depending on many factors, notably the reaction temperature and thenature of the reagents and solvent employed. However, provided that thereaction may be effected under the preferred conditions outlined above,a period of from 5 minutes to 48 hours, more preferably from 30 minutesto 24 hours, will usually suffice.

Step 1c

A compound of formula 1-4 can be prepared by reduction of a compound offormula 1-3 with a suitable reducing agent, such as sodium borohydride(NaBH₄), lithium aluminumhydride (LAH), diborane, hydrogen and a metalcatalyst, iron and hydrochoric acid, stannic chloride and hydrochoricacid, zinc and hydrochoric acid, formic acid, borane dimethylsulfidecomplex, borane-THF, (preferably hydrogen and a metal catalyst), usuallyin excess, in a reaction inert solvent such as methanol, ethanol,propanol, butanol, terahydrofuran (THF) (preferably methanol orethanol), generally at temperature of −78° C. to 60° C., preferably fromabout 0° C. to 45° C. for 5 minutes to 24 hours, preferably 60 minutesto 12 hours.

Step 1d

In step 1d, an amine compound of formula 1-4 can be prepared by thereductive amination of the alkanone compound with an amine compound offormula 1-5 in a similar condition in step 1a.

Alternatively, a compound of formula 1-4 can be prepared by alkylationof a compound of formula 1-5 with compound of formula Z—R¹.

The reaction can take place over a wide range of temperatures, and theprecise reaction temperature is not critical to the invention. Thepreferred reaction temperature will depend upon such factors as thenature of the solvent, and the starting material or reagent used.However, in general, it is convenient to carry out the reaction at atemperature of from 0° C. to 120° C., more preferably from 0° C. to 70°C. The time required for the reaction may also vary widely, depending onmany factors, notably the reaction temperature and the nature of thereagents and solvent employed. However, provided that the reaction maybe effected under the preferred conditions outlined above, a period offrom 5 minutes to 48 hours, more preferably from 30 minutes to 24 hours,will usually suffice.

Step 1e

A compound of formula 1-A can be prepared by cyclization of a compoundof formula 1-4 with a suitable carbonylating agent such ascarbonyldiimidazole, trichloromethyl chloroformate, triphosgene and urea(preferably carbonyldiimidazole), usually in excess, in a reaction inertsolvent such as dimethoxyethane, dioxane, acetonitrile,N,N′-dimethylformamide, dimethylsulfoxide, dichloromethane,dichloroethane, chloroform, or terahydrofuran (THF) (preferably THF),generally at temperature of −78° C. to 120° C., preferably from about20° C. to 100° C. for 5 minutes to 24 hours, preferably 60 minutes to 12hours.

Alternatively, the compound of 1-A (wherein R¹ is isopropyl as shown inScheme 1b) can be prepared from an alkenyl-benzimidazolone compound offormula 1-6 according to the following Scheme 1b in a reaction conditionknown to a skilled person.

Synthesis of Amine Moiety (2-A):

The following reaction Schemes illustrate the preparation of piperidinecompounds of formula (2-A).

In the above formulae, PG represents a protecting group. The term“protecting group”, as used herein, means an amino protecting groupwhich is selected from typical amino protecting groups described inProtective Groups in Organic Synthesis edited by T. W. Greene et al.(John Wiley & Sons, 1991). Typical amino protecting groups includebenzyl, C₂H_(S)O(C═O)—, CH₃(C═O)—, tert-butyldimethylsilyl (TBS),tert-butyldiphenylsilyl, benzyloxycarbonyl represented as Z andtert-buthoxycarbonyl represented as t-Boc or Boc.

A compound of formula 2-2 can be prepared by alkylation or reductiveamination of a compound of formula 2-1 with a compound of formulaalkyl-R³, halo-R³, or H(C═O)—R³ in a similar condition to step 1a. When—B—R³ represents 4-hydroxytetrahydropyranylmethyl, this alkylation canbe done by using a 1,6-dioxaspiro[2.5]octane compound.

Then, this reaction is followed by deprotection to obtain a compound offormula 1-A. This deprotection may be carried out according toprocedures known to those skilled in the art to give the compound offormula of 2-A.

Alternatively, the compound of formula (2-A) can be prepared from apiperidine compound of formula 2-3 according to the following Scheme 2bwith a reaction condition known to a skilled person.

For example, in step 2c, the compound 2-4 may be prepared by alkylationor reductive amination in essentially the same condition as onedescribed in step 2a of Scheme 2a. Then, the reduction in step 2d may becarried out in the presence of a reducing reagent such as LiAlH₄ in areaction inert solvent such as THF. Suitable reaction temperature rangesfrom about −78° C. to about 100° C., preferably from about −30° C. toabout 40° C.

The compound of formula (1-A) can be prepared from a piperidine compoundof formula 2-5 according to the following Scheme 2c with a reactioncondition known to a skilled person

For example, in step 2e, the compounds 2-6 may be prepared by alkylationor reductive amination in a similar condition to one described in step2a of scheme 2a. Then, the reduction in step 2f may be carried out inthe presence of a H₂ and a hydrogenation catalyst such as PtO₂ in areaction inert solvent such as THF. Suitable reaction temperature rangesfrom about −78° C. to about 100° C., preferably from about −30° C. toabout 40° C.

Synthesis of the Compound of Formula (I):

The following reaction Schemes illustrate the preparation ofbenzimidazolone compounds of formula I.

Step 3a:

A compound of formula 3-1 can be prepared by carbonylation of a compoundof formula 1-A with a compound of formula 2-A in the presence of asuitable carbonylating agent such as carbonyldiimidazole,trichloromethyl chloroformate, triphosgene, 4-nitrophenyl chloroformate,or urea (preferably triphosgene), usually in excess, in a reaction inertsolvent such as, dimethoxyethane, dioxane, acetonitrile,N,N′-dimethylformamide, dimethylsulfoxide, dichloromethane,dichloroethane, terahydrofuran (THF), benzene, toluene, or chloroform(preferably THF), generally at temperature of −78° C. to 120° C.,preferably from about 0° C. to 90° C. for 5 minutes to 24 hours,preferably 60 minutes to 12 hours.

Step 3b:

A compound of formula 3-2 is prepared by deprotection of a compound offormula 3-1 with an acid such as hydrochloride,

Step 3c:

A compound of formula (Ia) can be prepared by alkylation or reductiveamination in a similar condition to one described in step 2a of Scheme2a.

Alternatively, the compound of formula (la) can be prepared fromalkyl-benzimidazolone compounds according to the following Scheme 3b ina reaction condition known to a skilled person.

For example, in step 3d, the compound of formula 1-A can be reacted witha compound of formula 2-A in the presence of a carbonylating agent suchas carbonyldiimidazole, trichloromethyl chloroformate, triphosgene,4-nitrophenyl chloroformate, or urea (preferably triphosgene), usuallyin excess, in a reaction inert solvent such as dimethoxyethane, dioxane,acetonitrile, N,N′-dimethylformamide, dimethylsulfoxide,dichloromethane, dichloroethane, terahydrofuran (THF), benzene, toluene,or chloroform (preferably THF), generally at temperature of −78° C. to120° C., preferably from about 0° C. to 90° C. for 5 minutes to 24hours, preferably 60 minutes to 12 hours.

The compound of formula 7 can be prepared by using a reaction known to askilled person. For example, the compound of formula 7 can be preparedfrom a compound of formula 3 according to the following Scheme 3c in areaction condition known to a skilled person.

In the above Schemes from la to 3c, examples of suitable solventsinclude a mixture of any two or more of those solvents described in eachStep.

The compounds of formula (I), and the intermediates above-mentionedpreparation methods can be isolated and purified by conventionalprocedures, such as distillation, recrystallization or chromatographicpurification.

The optically active compounds of this invention can be prepared byseveral methods. For example, the optically active compounds of thisinvention may be obtained by chromatographic separation, enzymaticresolution or fractional crystallization from the final compounds.

Several compounds of this invention possess an asymmetric center. Hence,the compounds can exist in separated (+)- and (−)-optically activeforms, as well as in racemic one thereof. The present invention includesall such forms within its scope. Individual isomers can be obtained byknown methods, such as optically selective reaction or chromatographicseparation in the preparation of the final product or its intermediate.

The subject invention also includes isotopically-labelled compounds,which are identical to those recited in formula (I), but for the factthat one or more atoms are replaced by an atom having an atomic mass ormass number different from the atomic mass or mass number usually foundin nature. Examples of isotopes that can be incorporated into compoundsof the invention include isotopes of hydrogen, carbon, nitrogen, oxygen,phosphorus, fluorine and chlorine, such as ²H, ³H, ¹³C, ¹⁴C, ¹⁵N, ¹⁸O,¹⁷O, ³¹P, ³²P, ³⁵S, ¹⁸F, and ³⁶Cl, respectively. Compounds of thepresent invention, prodrugs thereof, pharmaceutically acceptable estersof said compounds and pharmaceutically acceptable salts of saidcompounds, of said esters or of said prodrugs which contain theaforementioned isotopes and/or other isotopes of other atoms are withinthe scope of this invention. Certain isotopically-labelled compounds ofthe present invention, for example those into which radioactive isotopessuch as ³H and ¹⁴C are incorporated, are useful in drug and/or substratetissue distribution assay. Tritiated, i.e., ³H, and carbon-14, i.e.,¹⁴C, isotopes are particularly preferred for their ease of presentationand detectability. Further, substitution with heavier isotopes such asdeuterium, i.e., ²H, can afford therapeutic advantage resulting fromgreater metabolic stability, for example increased in vivo half-life orreduced dosage requirement and, hence, may be preferred in somecircumstances. Isotopically labeled compounds of formula (I) of thisinvention and prodrugs thereof can generally be prepared by carrying outthe procedure disclosed in above-disclosed Schemes and/or Examples andPreparations below, by submitting a readily available isotopicallylabeled reagent for a non-isotopically labeled reagent.

The present invention includes salt forms of the compounds (I) asobtained.

Pharmaceutically acceptable salts of the compounds of formula (I)include the acid addition and base addition salts (including disalts)thereof.

Pharmaceutically acceptable non-toxic salts of compounds of formula (I)may be prepared by conventional techniques by, for example, contactingsaid compound with a stoichiometric amount of an appropriate alkali oralkaline earth metal (sodium, potassium, calcium and magnesium)hydroxide or alkoxide in water or an appropriate organic solvent such asethanol, isopropanol, mixtures thereof, or the like.

The bases which are used to prepare the pharmaceutically acceptable baseaddition salts of the acidic compounds of this invention of formula (I)are those which form non-toxic base addition salts, i.e., saltscontaining pharmaceutically acceptable cations, such as adenine,arginine, cytosine, lysine, benethamine (i.e.,N-benzyl-2-phenyletylamine), benzathine (i.e.,N,N-dibenzylethylenediamine), choline, diolamine (i.e., diethanolamine),ethylenediamine, glucosamine, glycine, guanidine, guanine, meglumine(i.e., N-methylglucamine), nicotinamide, olamine (i.e., ethanolamine),ornithine, procaine, proline, pyridoxine, serine, tyrosine, valine andtromethamine (i.e., tris or tris(hydroxymethyl)aminomethane). The baseaddition salts can be prepared by conventional procedures.

Insofar as the certain compounds of this invention are basic compounds,they are capable of forming a wide variety of different salts withvarious inorganic and organic acids.

The acids which are used to prepare the pharmaceutically acceptable acidaddition salts of the basic compounds of this invention of formula (I)are those which form non-toxic acid addition salts, i.e., saltscontaining pharmaceutically acceptable anions, such as the chloride,bromide, iodide, nitrate, sulfate or bisulfate, phosphate or acidphosphate, acetate, lactate, citrate or acid citrate, tartrate orbi-tartrate, succinate, malate, fumarate, gluconate, saccharate,benzoate, methanesulfonate, ethanesulfonate, benzenesulfonate,p-toluenesulfonate, adipate, aspartate camsylate, edisylate (i.e.,1,2-ethanedisulfonate), estolate (i.e., laurylsulfate), gluceptate(i.e., gluscoheptonate), gluconate, 3-hydroxy-2-naphthoate, xionofoate(i.e., 1-hydrroxy-2-naphthoate), isethionate, (i.e.,2-hydroxyethanesulfonate), mucate (i.e., galactarate), 2-naphsylate(i.e., naphthalenesulphonate, stearate, cholate, glucuronate, glutamate,hippurate, lactobionate, lysinate, maleate, mandelate, napadisylate,nicatinate, polygalacturonate, salicylate, sulphosalicylate, tannate,tryptophanate, borate, carbonate, oleate, phthalate and pamoate (i.e.,1.1′-methylene-bis-(2-hydroxy-3-naphthoate). Of these, we preferedisylate (including hemi-edisylate) and hydrochloride. The acidaddition salts can be prepared by conventional procedures.

For a review of on suitable salts see Berge et al., J. Pharm. Sci., 66,1-19, 1977.

The present invention includes salt forms of the compounds of formula(2-A′) as obtained.

Compounds of formula (2-A′) may be capable of forming cations. Cationsof compounds of formula (2-A′) may be prepared by conventionaltechniques by, for example, contacting said compound with astoichiometric amount of an appropriate alkali or alkaline earth metal(sodium, potassium, calcium and magnesium) hydroxide or alkoxide inwater or an appropriate organic solvent such as ethanol, isopropanol,mixtures thereof, or the like.

The bases used to prepare the base addition salts of the acidiccompounds of formula (2-A′) are those which form base addition salts.Such base addition salts include pharmaceutically acceptable baseaddition salts as described above and salts containing cations, such as.triethylamine, pyridine and ammonia.

The compounds of formula (2-A′) are capable of forming a wide variety ofdifferent salts with various inorganic and organic acids.

The acids used to prepare the acid addition salts of the compound offormula (2-A′) are those which form acid addition salts. Such acidaddition salts include pharmaceutically acceptable acid addition saltsas described above and salts containing anions, such as cyanide.

Also included within the scope of this invention are bioprecursors (alsocalled pro-drugs) of the compounds of the formula (I). A bioprecursor ofa compound of the formula (I) is a chemical derivative thereof which isreadily converted back into the parent compound of the formula (I) inbiological systems. In particular, a bioprecursor of a compound of theformula (I) is converted back to the parent compound of the formula (I)after the bioprecursor has been administered to, and absorbed by, amammalian subject, e.g., a human subject. For example, it is possible tomake a bioprecursor of the compounds of formula (I) in which one or bothof L and W include hydroxy groups by making an ester of the hydroxygroup. When only one of L and W includes hydroxy group, only mono-esteris possible. When both L and W include hydroxy, mono- and di-esters(which can be the same or different) can be made. Typical esters aresimple alkanoate esters, such as acetate, propionate, butyrate, etc. Inaddition, when L or W includes a hydroxy group, bioprecursors can bemade by converting the hydroxy group to an acyloxymethyl derivative(e.g., a pivaloyloxymethyl derivative) by reaction with an acyloxymethylhalide (e.g., pivaloyloxymethyl chloride).

When the compounds of the formula (I) of this invention may formsolvates such as hydrates, such solvates are included within the scopeof this invention.

Compounds of formula (I) containing one or more asymmetric carbon atomscan exist as two or more stereoisomers. Where a compound of formula (I)contains an alkenyl or alkenylene group, geometric cis/trans (or Z/E)isomers are possible. Where the compound contains, for example, a ketoor oxime group or an aromatic moiety, tautomeric isomerism(‘tautomerism’) can occur. It follows that a single compound may exhibitmore than one type of isomerism.

Included within the scope of the present invention are allstereoisomers, geometric isomers and tautomeric forms of the compoundsof formula (I), including compounds exhibiting more than one type ofisomerism, and mixtures of one or more thereof. Also included are acidaddition or base salts wherein the counterion is optically active, forexample, D-lactate or L-lysine, or racemic, for example, DL-tartrate orDL-arginine.

Cis/trans isomers may be separated by conventional techniques well knownto those skilled in the art, for example, chromatography and fractionalcrystallisation.

Conventional techniques for the preparation/isolation of individualenantiomers include chiral synthesis from a suitable optically pureprecursor or resolution of the racemate (or the racemate of a salt orderivative) using, for example, chiral high pressure liquidchromatography (HPLC).

Alternatively, the racemate (or a racemic precursor) may be reacted witha suitable optically active compound, for example, an alcohol, or, inthe case where the compound of formula (I) contains an acidic or basicmoiety, an acid or base such as tartaric acid or 1-phenylethylamine. Theresulting diastereomeric mixture may be separated by chromatographyand/or fractional crystallization and one or both of thediastereoisomers converted to the corresponding pure enantiomer(s) bymeans well known to a skilled person.

Stereoisomeric conglomerates may be separated by conventional techniquesknown to those skilled in the art—see, for example, “Stereochemistry ofOrganic Compounds” by E L Eliel (Wiley, New York, 1994).

Compounds of the invention intended for pharmaceutical use may beadministered as crystalline or amorphous products. They may be obtained,for example, as solid plugs, powders, or films by methods such asprecipitation, crystallization, freeze drying, spray drying, orevaporative drying. Microwave or radio frequency drying may be used forthis purpose.

They may be administered alone or in combination with one or more othercompounds of the invention or in combination with one or more otherdrugs (or as any combination thereof). Generally, they will beadministered as a formulation in association with one or morepharmaceutically acceptable excipients. The term “excipient” is usedherein to describe any ingredient other than the compound(s) of theinvention. The choice of excipient will to a large extent depend onfactors such as the particular mode of administration, the effect of theexcipient on solubility and stability, and the nature of the dosageform.

Pharmaceutical compositions suitable for the delivery of compounds ofthe present invention and methods for their preparation will be readilyapparent to those skilled in the art. Such compositions and methods fortheir preparation may be found, for example, in ‘Remington'sPharmaceutical Sciences’, 19th Edition (Mack Publishing Company, 1995).

Oral Administration

The compounds of the invention may be administered orally. Oraladministration may involve swallowing, so that the compound enters thegastrointestinal tract, or buccal or sublingual administration may beemployed by which the compound enters the blood stream directly from themouth.

Formulations suitable for oral administration include solid formulationssuch as tablets, capsules containing particulates, liquids, or powders,lozenges (including liquid-filled), chews, multi- and nano-particulates,gels, solid solution, liposome, films (including muco-adhesive), ovules,sprays and liquid formulations.

Liquid formulations include suspensions, solutions, syrups and elixirs.Such formulations may be employed as fillers in soft or hard capsulesand typically comprise a carrier, for example, water, ethanol,polyethylene glycol, propylene glycol, methylcellulose, or a suitableoil, and one or more emulsifying agents and/or suspending agents. Liquidformulations may also be prepared by the reconstitution of a solid, forexample, from a sachet.

The compounds of the invention may also be used in fast-dissolving,fast-disintegrating dosage forms such as those described in ExpertOpinion in Therapeutic Patents, 11 (6), 981-986 by Liang and Chen(2001).

For tablet dosage forms, depending on dose, the drug may make up from 1wt % to 80 wt % of the dosage form, more typically from 5 wt % to 60 wt% of the dosage form. In addition to the drug, tablets generally containa disintegrant. Examples of disintegrants include sodium starchglycolate, sodium carboxymethyl cellulose, calcium carboxymethylcellulose, croscannellose sodium, crospovidone, polyvinylpyrrolidone,methyl cellulose, microcrystalline cellulose, lower alkyl-substitutedhydroxypropyl cellulose, starch, pregelatinised starch and sodiumalginate. Generally, the disintegrant will comprise from 1 wt % to 25 wt%, preferably from 5 wt % to 20 wt % of the dosage form.

Binders are generally used to impart cohesive qualities to a tabletformulation. Suitable binders include microcrystalline cellulose,gelatin, sugars, polyethylene glycol, natural and synthetic gums,polyvinylpyrrolidone, pregelatinised starch, hydroxypropyl cellulose andhydroxypropyl methylcellulose. Tablets may also contain diluents, suchas lactose (monohydrate, spray-dried monohydrate, anhydrous and thelike), mannitol, xylitol, dextrose, sucrose, sorbitol, microcrystallinecellulose, starch and dibasic calcium phosphate dihydrate.

Tablets may also optionally comprise surface active agents, such assodium lauryl sulfate and polysorbate 80, and glidants such as silicondioxide and talc. When present, surface active agents may comprise from0.2 wt % to 5 wt % of the tablet, and glidants may comprise from 0.2 wt% to 1 wt % of the tablet.

Tablets also generally contain lubricants such as magnesium stearate,calcium stearate, zinc stearate, sodium stearyl fumarate, and mixturesof magnesium stearate with sodium lauryl sulphate. Lubricants generallycomprise from 0.25 wt % to 10 wt %, preferably from 0.5 wt % to 3 wt %of the tablet.

Other possible ingredients include anti-oxidants, colourants, flavouringagents, preservatives and taste-masking agents.

Exemplary tablets contain up to about 80% drug, from about 10 wt % toabout 90 wt % binder, from about 0 wt % to about 85 wt % diluent, fromabout 2 wt % to about 10 wt % disintegrant, and from about 0.25 wt % toabout 10 wt % lubricant.

Tablet blends may be compressed directly or by roller to form tablets.Tablet blends or portions of blends may alternatively be wet-, dry-, ormelt-granulated, melt congealed, or extruded before tabletting. Thefinal formulation may comprise one or more layers and may be coated oruncoated; it may even be encapsulated.

The formulation of tablets is discussed in “Pharmaceutical Dosage Forms:Tablets, Vol. 1”, by H. Lieberman and L. Lachman, Marcel Dekker, N.Y.,N.Y., 1980 (ISBN 0-8247-6918-X).

Solid formulations for oral administration may be formulated to beimmediate and/or modified release. Modified release formulations includedelayed-, sustained-, pulsed-, controlled-, targeted and programmedrelease.

Suitable modified release formulations for the purposes of the inventionare described in U.S. Pat. No. 6,106,864. Details of other suitablerelease technologies such as high energy dispersions and osmotic andcoated particles are to be found in Verma et al, PharmaceuticalTechnology On-line, 25(2), 1-14 (2001). The use of chewing gum toachieve controlled release is described in WO 00/35298.

Parental Administration

The compounds of the invention may also be administered directly intothe blood stream, into muscle, or into an internal organ. Suitable meansfor parenteral administration include intravenous, intraarterial,intraperitoneal, intrathecal, intraventricular, intraurethral,intrastemal, intracranial, intramuscular and subcutaneous. Suitabledevices for parenteral administration include needle (includingmicroneedle) injectors, needle-free injectors and infusion techniques.

Parenteral formulations are typically aqueous solutions which maycontain excipients such as salts, carbohydrates and buffering agents(preferably to a pH of from 3 to 9), but, for some applications, theymay be more suitably formulated as a sterile non-aqueous solution or asa dried form to be used in conjunction with a suitable vehicle such assterile, pyrogen-free water.

The preparation of parenteral formulations under sterile conditions, forexample, by lyophilisation, may readily be accomplished using standardpharmaceutical techniques well known to those skilled in the art.

The solubility of compounds of formula (I) used in the preparation ofparenteral solutions may be increased by the use of appropriateformulation techniques, such as the incorporation ofsolubility-enhancing agents.

Formulations for parenteral administration may be formulated to beimmediate and/or modified release. Modified release formulations includedelayed-, sustained-, pulsed-, controlled-, targeted and programmedrelease. Thus compounds of the invention may be formulated as a solid,semi-solid, or thixotropic liquid for administration as an implanteddepot providing modified release of the active compound. Examples ofsuch formulations include drug-coated stents and PGLA microspheres.

Topical Administration

The compounds of the invention may also be administered topically to theskin or mucosa, that is, dermally or transdermally. Typical formulationsfor this purpose include gels, hydrogels, lotions, solutions, creams,ointments, dusting powders, dressings, foams, films, skin patches,wafers, implants, sponges, fibres, bandages and microemulsions.Liposomes may also be used. Typical carriers include alcohol, water,mineral oil, liquid petrolatum, white petrolatum, glycerin, polyethyleneglycol and propylene glycol. Penetration enhancers may beincorporated—see, for example, J Pharm Sci, 88 (10), 955-958 by Finninand Morgan (October 1999).

Other means of topical administration include delivery byelectroporation, iontophoresis, phonophoresis, sonophoresis andmicroneedle or needle-free (e.g. Powderject™, Bioject™, etc.) injection.

Formulations for topical administration may be formulated to beimmediate and/or modified release. Modified release formulations includedelayed-, sustained-, pulsed-, controlled-, targeted and programmedrelease.

Inhaled/Intranasal Administration

The compounds of the invention can also be administered intranasally orby inhalation, typically in the form of a dry powder (either alone, as amixture, for example, in a dry blend with lactose, or as a mixedcomponent particle, for example, mixed with phospholipids, such asphosphatidylcholine) from a dry powder inhaler or as an aerosol sprayfrom a pressurised container, pump, spray, atomiser (preferably anatomiser using electrohydrodynamics to produce a fine mist), ornebuliser, with or without the use of a suitable propellant, such as1,1,1,2-tetrafluoroethane or 1,1,1,2,3,3,3-heptafluoropropane. Forintranasal use, the powder may comprise a bioadhesive agent, forexample, chitosan or cyclodextrin.

The pressurised container, pump, spray, atomizer, or nebuliser containsa solution or suspension of the compound(s) of the invention comprising,for example, ethanol, aqueous ethanol, or a suitable alternative agentfor dispersing, solubilising, or extending release of the active, apropellant(s) as solvent and an optional surfactant, such as sorbitantrioleate, oleic acid, or an oligolactic acid.

Prior to use in a dry powder or suspension formulation, the drug productis micronised to a size suitable for delivery by inhalation (typicallyless than 5 microns). This may be achieved by any appropriatecomminuting method, such as spiral jet milling, fluid bed jet milling,supercritical fluid processing to form nanoparticles, high pressurehomogenisation, or spray drying.

Capsules (made, for example, from gelatin or HPMC), blisters andcartridges for use in an inhaler or insufflator may be formulated tocontain a powder mix of the compound of the invention, a suitable powderbase such as lactose or starch and a performance modifier such asl-leucine, mannitol, or magnesium stearate. The lactose may be anhydrousor in the form of the monohydrate, preferably the latter. Other suitableexcipients include dextran, glucose, maltose, sorbitol, xylitol,fructose, sucrose and trehalose.

A suitable solution formulation for use in an atomiser usingelectrohydrodynamics to produce a fine mist may contain from 1 μg to 20mg of the compound of the invention per actuation and the actuationvolume may vary from 1 μl to 100 μl. A typical formulation may comprisea compound of formula (I), propylene glycol, sterile water, ethanol andsodium chloride. Alternative solvents which may be used instead ofpropylene glycol include glycerol and polyethylene glycol.

Suitable flavours, such as menthol and levomenthol, or sweeteners, suchas saccharin or saccharin sodium, may be added to those formulations ofthe invention intended for inhaled/intranasal administration.

Formulations for inhaled/intranasal administration may be formulated tobe immediate and/or modified release using, for example,poly(DL-lactic-coglycolic acid (PGLA). Modified release formulationsinclude delayed-, sustained-, pulsed-, controlled-, targeted andprogrammed release.

In the case of dry powder inhalers and aerosols, the dosage unit isdetermined by means of a valve which delivers a metered amount. Units inaccordance with the invention are typically arranged to administer ametered dose or “puff” containing from 1 to 100 μg of the compound offormula (I). The overall daily dose will typically be in the range 50 μgto 20 mg which may be administered in a single dose or, more usually, asdivided doses throughout the day.

Rectal/Intravaginal Administration

The compounds of the invention may be administered rectally orvaginally, for example, in the form of a suppository, pessary, or enema.Cocoa butter is a traditional suppository base, but various alternativesmay be used as appropriate.

Formulations for rectal/vaginal administration may be formulated to beimmediate and/or modified release. Modified release formulations includedelayed-, sustained-, pulsed-, controlled-, targeted and programmedrelease.

Ocular/Aural Administration

The compounds of the invention may also be administered directly to theeye or ear, typically in the form of drops of a micronised suspension orsolution in isotonic, pH-adjusted, sterile saline. Other formulationssuitable for ocular and aural administration include ointments,biodegradable (e.g. absorbable gel sponges, collagen) andnon-biodegradable (e.g. silicone) implants, wafers, lenses andparticulate or vesicular systems, such as niosomes or liposomes. Apolymer such as crossed-linked polyacrylic acid, polyvinylalcohol,hyaluronic acid, a cellulosic polymer, for example,hydroxypropylmethylcellulose, hydroxyethylcellulose, or methylcellulose, or a heteropolysaccharide polymer, for example, gelan gum,may be incorporated together with a preservative, such as benzalkoniumchloride. Such formulations may also be delivered by iontophoresis.

Formulations for ocular/aural administration may be formulated to beimmediate and/or modified release. Modified release formulations includedelayed-, sustained-, pulsed-, controlled-, targeted, or programmedrelease.

Other Technologies

The compounds of the invention may be combined with solublemacromolecular entities, such as cyclodextrin and suitable derivativesthereof or polyethylene glycol-containing polymers, in order to improvetheir solubility, dissolution rate, taste-masking, bioavailabilityand/or stability for use in any of the aforementioned modes ofadministration.

Drug-cyclodextrin complexes, for example, are found to be generallyuseful for most dosage forms and administration routes. Both inclusionand non-inclusion complexes may be used. As an alternative to directcomplexation with the drug, the cyclodextrin may be used as an auxiliaryadditive, i.e. as a carrier, diluent, or solubiliser. Most commonly usedfor these purposes are alpha-, beta- and gamma-cyclodextrins, examplesof which may be found in International Patent Applications Nos. WO91/11172, WO 94/02518 and WO 98/55148.

Kit-of-Parts

Inasmuch as it may desirable to administer a combination of activecompounds, for example, for the purpose of treating a particular diseaseor condition, it is within the scope of the present invention that twoor more pharmaceutical compositions, at least one of which contains acompound in accordance with the invention, may conveniently be combinedin the form of a kit suitable for coadministration of the compositions.

Thus the kit of the invention comprises two or more separatepharmaceutical compositions, at least one of which contains a compoundof formula (I) in accordance with the invention, and means forseparately retaining said compositions, such as a container, dividedbottle, or divided foil packet. An example of such a kit is the familiarblister pack used for the packaging of tablets, capsules and the like.

The kit of the invention is particularly suitable for administeringdifferent dosage forms, for example, oral and parenteral, foradministering the separate compositions at different dosage intervals,or for titrating the separate compositions against one another. Toassist compliance, the kit typically comprises directions foradministration and may be provided with a so-called memory aid.

Dosage

For administration to human patients, the total daily dose of thecompounds of the invention is typically in the range 0.05 mg to 100 mgdepending, of course, on the mode of administration, preferred in therange 0.1 mg to 50 mg and more preferred in the range 0.5 mg to 20 mg.For example, oral administration may require a total daily dose of from1 mg to 20 mg, while an intravenous dose may only require from 0.5 mg to10 mg. The total daily dose may be administered in single or divideddoses.

These dosages are based on an average human subject having a weight ofabout 65 kg to 70 kg. The physician will readily be able to determinedoses for subjects whose weight falls outside this range, such asinfants and the elderly.

For the avoidance of doubt, references herein to “treatment” includereferences to curative, palliative and prophylactic treatment.

A 5-HT₄ agonist of the present invention may be usefully combined withanother pharmacologically active compound, or with two or more otherpharmacologically active compounds, particularly in the treatment ofgastroesophageal reflux disease. For example, a 5-HT₄ agonist,particularly a compound of the formula (I), or a pharmaceuticallyacceptable salt or solvate thereof, as defined above, may beadministered simultaneously, sequentially or separately in combinationwith one or more agents selected from:

-   (i) histamine H₂ receptor antagonists, e.g. ranitidine, lafutidine,    nizatidine, cimetidine, famotidine and roxatidine;-   (ii) proton pump inhibitors, e.g. omeprazole, esomeprazole,    pantoprazole, rabeprazole, tenatoprazole, ilaprazole and    lansoprazole;-   (iii) Acid pump antagonists, e.g. soraprazan, revaprazan(YH-1885),    AZD-0865, CS-526, AU-2064 and YJA-20379-8;-   (iv) oral antacid mixtures, e.g. Maalox®, Aludrox® and Gaviscon®;-   (v) mucosal protective agents, e.g. polaprezinc, ecabet sodium,    rebamipide, teprenone, cetraxate, sucralfate, chloropylline-copper    and plaunotol;-   (vi) GABA_(B) agonists, e.g. baclofen and AZD-3355;-   (vii) α2 agonists, e.g. clonidine, medetomidine, lofexidine,    moxonidine, tizanidine, guanfacine, guanabuz, talipexole and    dexmedetomidine;-   (viii) Xanthin derivatives, e.g. Theophylline, aminophylline and    doxofylline;-   (ix) calcium channel blockers, e.g. aranidipine, lacidipine,    falodipine, azelnidipine, clinidipine, lomerizine, diltiazem,    gallopamil, efonidipine, nisoldipine, amlodipine, lercanidipine,    bevantolol, nicardipine, isradipine, benidipine, verapamil,    nitrendipine, bamidipine, propafenone, manidipine, bepridil,    nifedipine, nilvadipine, nimodipine, nifedipine and fasudil;-   (x) benzodiazepine agonists, e.g. diazepam, zaleplon, zolpidem,    haloxazolam, clonazepam, prazepam, quazepam, flutazolam, triazolam,    lormetazepam, midazolam, tofisopam, clobazam, flunitrazepam and    flutoprazepam;-   (xi) prostaglandin analogues, e.g. Prostaglandin, misoprostol,    treprostinil, esoprostenol, latanoprost, iloprost, beraprost,    enprostil, ibudilast and ozagrel;-   (xii) histamine H₃ agonists, e.g. R-alpha-methylhistamine and    BP-294;-   (xiii) anti-gastric agents, e.g. Anti-gastrin vaccine, itriglumide    and Z-360;-   (xiv) 5-HT₃ antagonists, e.g. dolasetron, palonosetron, alosetron,    azasetron, ramosetron, mitrazapine, granisetron, tropisetron,    E-3620, ondansetron and indisetron;-   (xv) tricyclic antidepressants, e.g. imipramine, amitriptyline,    clomipramine, amoxapine and lofepramine;-   (xvi) GABA agonists, e.g. gabapentin, topiramate, cinolazepam,    clonazepam, progabide, brotizolam, zopiclone, pregabalin and    eszopiclone;-   (xvii) opioid analgesics, e.g. morphine, heroin, hydromorphone,    oxymorphone, levorphanol, levallorphan, methadone, meperidine,    fentanyl, cocaine, codeine, dihydrocodeine, oxycodone, hydrocodone,    propoxyphene, nalmefene, nalorphine, naloxone, naltrexone,    buprenorphine, butorphanol, nalbuphine and pentazocine;-   (xviii) somatostatin analogues, e.g. octreotide, AN-238 and    PTR-3173;-   (xix) Cl Channel activator: e.g. lubiprostone;-   (xx) selective serotonin reuptake inhibitors, e.g. sertraline,    escitalopram, fluoxetine, nefazodone, fluvoxamine, citalopram,    milnacipran, paroxetine, venlafaxine, tramadol, sibutramine,    duloxetine, desvenlafaxine and depocxetine;-   (xxi) anticholinergics, e.g. dicyclomine and hyoscyamine;-   (xxii) laxatives, e.g. Trifyba®, Fybogel®, Konsyl®, Isogel®,    Regulan®, Celevac® and Normacol®;-   (xxiii) fiber products, e.g. Metamucil®;-   (xxiv) antispasmodics, e.g.: mebeverine;-   (xxv) dopamine antagonists, e.g. metoclopramide, domperidone and    levosulpiride;-   (xxvi) cholinergics, e.g. neostigmine-   (xxvii) AChE inhibitor: galantamine, metrifonate, rivastigmine,    itopride and donepezil;-   (xxvi ii) Tachykinin (NK) antagonists, particularly NK-3, NK-2 and    NK-1 e.g.

antagonists, nepadutant, saredutant, talnetant,(αR,9R)-7-[3,5-bis(trifluoromethyl)benzyl]-8,9,10,11-tetrahydro-9-methyl-5-(4-methylphenyl)-7H-[1,4]diazocino[2,1-g][1,7]naphthridine-6-13-dione(TAK-637),5-[[(2R,3S)-2-[(1R)-1-[3,5-bis(trifluoromethyl)phenyl]ethoxy-3-(4-fluorophenyl)-4-morpholinyl]methyl]-1,2-dihydro-3H-1,2,4-triazol-3-one(MK-869), lanepitant, dapitant and3-[[2-methoxy-5-(trifluoromethoxy)phenyl]methylamino]-2-phenyl-piperidine(2S,3S).

Method for Assessing Biological Activities:

The 5-HT₄ receptor binding affinities of the compounds of this inventionare determined by the following procedures.

Membrane Preparation

Pig heads were supplied from an abattoir. Striatal tissues weredissected, weighed and homogenized in 15 volumes of 50 mM ice-cold HEPES(pH 7.5) in a Polytron homogenizer (30 sec at full speed). Suspensionwas centrifuged at 48,000 g and 4° C. for 15 min. The resulting pelletwas resuspended in an appropriate volume of 50 mM ice-cold HEPES,dispensed into aliquots and stored at −80° C. until use.

Bovine heads were also supplied from an abattoir. Striatal tissues weredissected, weighed and homogenized in 20 volumes of 50 mM ice-coldTris-HCl (pH 7.4) in a Polytron homogenizer (30 sec at full speed).Suspension was centrifuged at 20,000 g and 4° C. for 30 min. Theresulting pellet was resuspended in 15 volumes of 50 mM ice-coldTris-HCl, homegenized and centrifuged again in the same way. The finalpellet was resuspended in an appropriate volume of 50 mM Tris-HCl,dispensed into aliquots and stored at −80° C. until use.

Cerebral cortical tissues were removed from male Sprague-Dawley (SD)rats (Japan SLC), weighed and placed in 10 volumes of 50 mM ice-coldTris-HCl (pH 7.5). This was homogenized in a Polytron homogenizer (30sec at full speed) and subsequently centrifuged at 48,000 g and 4° C.for 15 min. The resulting pellet was resuspended in 50 mM ice-coldTris-HCl, homegenized and centrifuged again in the same way. The finalpellet was resuspended in an appropriate volume of 50 mM Tris-HCl,dispensed into aliquots and stored at −80° C. until use.

The protein concentrations of homogenates were determined by Bradfordmethod or BCA protein method (Pierce) with BSA as a standard.

Binding Assays

Affinity of compounds for pig or bovine 5-HT₄ and rat 5-HT₃ receptorswere assessed with using radiolabeled specific ligands, GR 113808({1-[2-(methylsulfonyl)ethyl]-4-piperidinyl}[methyl-3H]-1H-indole-3-carboxylate)and BRL 43694(1-Methyl-N-(9-[methyl-³H]-9-azabicyclo[3.3.1]non-3-yl)-1H-indazole-3-caboxamide).Compounds were incubated with 25-100 pM of [³H]-GR 113808 (Amersham) and0.6-1 mg protein of pig or bovine striatal membranes suspended in afinal volume of 0.8-1 ml of 50 mM Tris-HCl (pH 7.5). Nonspecific bindingwas determined with 10-50 μM 5-HT. The binding of 0.3 nM [³H]-BRL 43694(NEN) was measured using 400 μg protein of rat cortical membranessuspended in a final volume of 500 μl of 50 mM Tris-HCl (pH 7.5).Nonspecific binding was determined with 10 μM 5-HT.

The plates were incubated at room temperature on a plate shaker for 30min. The assays were stopped by rapid filtration using a Brandell cellharvester through Wallac-B filters pre-soaked in 0.2% poly(ethylenimine)at 4° C. for 60-90min. The filters were washed three times with 1 ml ofice-cold 50 mM HEPES, and were dried in a microwave or at roomtemperature. They were bagged and heated with meltilex scintillant(Wallac) or soaked in BetaplateScint (Wallac). Receptor-boundradioactivity was quantified using Big-spot counter, Betaplate counter(Wallac) or LS counter (Packard).

Human 5-HT₄ Binding(1)

Human 5-HT_(4(d)) transfected HEK293 cells were prepared and grownin-house. The collected cells were suspended in 50 mM HEPES (pH 7.4 at4° C.) supplemented with protease inhibitor cocktail (Boehringer, 1:1000dilution) and homogenized using a hand held Polytron PT 1200 disruptorset at full power for 30 sec on ice. The homogenates were centrifuged at40,000×g at 4° C. for 30 min. The pellets were then resuspended in 50 mMHEPES (pH 7.4 at 4° C.) and centrifuged once more in the same manner.The final pellets were resuspended in an appropriate volume of 50 mMHEPES (pH 7.4 at 25° C.), homogenized, aliquoted and stored at −80° C.until use. An aliquot of membrane fractions was used for proteinconcentration determination using BCA protein assay kit (PIERCE) andARVOsx plate reader (Wallac).

For the binding experiments, 25 μl of test compounds were incubated with25 μl of [³H]-GR113808 (Amersham, final 0.2 nM) and 150 μl of membranehomogenate and WGA-SPA beads (Amersham) suspension solutions (10 μgprotein and 1 mg SPA beads/well) for 60 minutes at room temperature.Nonspecific binding was determined by 1 μM GR113808 (Tocris) at thefinal concentration. Incubation was terminated by centrifugation at 1000rpm. Receptor-bound radioactivity was quantified by counting withMicroBeta plate counter (Wallac).

All compounds prepared in the working examples as described below weretested by this method, and they showed Ki values from 0.3 nM to 30 nMwith respect to inhibition of binding at the 5-HT₄ receptor.

Human 5-HT₄ Binding(2)

Human 5-HT₄(_(d)) transfected HEK293 cells were prepared and grownin-house. The collected cells were suspended in 50 mM Tris buffer (pH7.4 at 4° C.) supplemented with protease inhibitor cocktail (Boehringer,1:1000 dilution) and homogenized using a hand held Polytron PT 1200disruptor set at full power for 30 sec on ice. The homogenates werecentrifuged at 40,000×g at 4° C. for 10 min. The pellets were thenresuspended in 50 mM Tris buffer (pH 7.4 at 4° C.) and centrifuged oncemore in the same manner. The final pellets were resuspended in anappropriate volume of 50 mM Tris buffer (pH 7.4 at 25° C.) containing 10mM MgCl₂, homogenized, aliquoted and stored at −80° C. until use. Analiquot of membrane fractions was used for protein concentrationdetermination using BCA protein assay kit (PIERCE) and ARVOsx platereader (Wallac).

For the binding experiments, 50 μl of test compounds were incubated with50 μl of [³H]5-HT (Amersham, final 8.0 nM) and 400 μl of membranehomogenate (300 μg protein/tube) for 60 minutes at room temperature.Nonspecific binding was determined by 50 μM GR113808 (Tocris) at thefinal concentration. All incubations were terminated by rapid vacuumfiltration over 0.2% PEI soaked glass fiber filter papers using BRANDELharvester followed by three washes with 50 mM Tris buffer (pH 7.4 at 25°C.). Receptor-bound radioactivity was quantified by liquid scintillationcounting using Packard LS counter.

All compounds of Examples showed 5HT₄ receptor affinity.

Functional Assay:

The presence of 5-HT₄ receptors in the rat oesophagus and the ability todemonstrate partial agonism in the TMM preparation are reported in theliterature (See G. S. Baxter et al. Naunyn-Schmiedeberg's Arch Pharmacol(1991) 343: 439-446; M. Yukiko et al. JPET (1997) 283: 1000-1008; and J.J. Reeves et al. Br. J. Pharmacol. (1991) 103: 1067-1072). Morespecifically, partial agonist activity can be measured according to thefollowing procedures.

Male SD rats (Charles River) weighing 250-350 g were stunned and thenkilled by cervical dislocation. The oesophagus was dissected fromimmediately proximal to the stomach (including piece of stomach to markdistal end) up to the level of the trachea and then placed in freshKrebs' solution.

The outer skeletal muscle layer was removed in one go by peeling it awayfrom the underlying smooth muscle layer using forceps (stomach totracheal direction). The remaining inner tube of smooth muscle was knownas the TMM. This was trimmed to 2 cm from the original ‘stomach-end’ andthe rest discarded.

The TMMs were mounted as whole ‘open’ tubes in longitudinal orientationin 5 ml organ baths filled with warm (32° C.) aerated Krebs. Tissueswere placed under an initial tension of 750 mg and allowed toequilibrate for 60 minutes. The tissues were re-tensioned twice at 15minute intervals during the equilibration period. The pump flow rate wasset to 2 ml/min during this time.

Following equilibration, the pump was switched off. The tissues wereexposed to 1 μM carbachol and contracted and reached a steadycontractile plateau within 15 minutes. Tissues were then subject to 1 μM5-HT (this was to prime the tissues). The tissues relaxed in response to5-HT fairly rapidly—within 1 minute. As soon as maximal relaxation hasoccurred and a measurement taken, the tissues were washed at maximumrate (66 ml/min) for at least 1 minute and until the original baseline(pre-carbachol and 5-HT) has returned (usually, the baseline drops belowthe original one following initial equilibration). The pump flow ratewas reduced to 2 mL/min and the tissues left for 60 minutes.

A cumulative concentration-effect-curve (CEC) to 5-HT was constructedacross the range 0.1 nM to 1 μM, in half-log unit increments (5-HT curve1 for data analysis). Contact time between doses was 3 minutes or untilplateau established. Tissues responded quicker as concentration of 5-HTin the bath increases. At the end of the curve, the tissues were washed(at maximum rate) as soon as possible to avoid desensitisation ofreceptors. Pump rate was reduced to 2 ml/min and the tissues left for 60minutes.

A second CEC was carried out—either to 5-HT (for time control tissues),another 5-HT₄ agonist (standard) or a test compound (curve 2 for dataanalysis). Contact time varied for other 5-HT₄ agonists and testcompounds and was tailored according to the tissues' individualresponses to each particular agent. In tissues exposed to a testcompound, a high concentration (1 μM) of a 5-HT₄ antagonist (SB 203,186:1H-Indole-3-carboxylic acid, 2-(1-piperidinyl)ethyl ester, Tocris) wasadded to the bath following the last concentration of test compound.This was to see if any agonist-induced relaxation (if present) could bereversed. SB 203,186 reversed 5-HT induced relaxation, restoring thetissue's original degree of carbachol-induced tone.

Agonist activity of test compounds was confirmed by pre-incubatingtissues with 100 nM standard 5HT₄ antagonist such as SB 203,186. SB203,186 was added to the bath 5 minutes before the addition of carbacholprior to curve 2. Tissues must be ‘paired’ for data analysis i.e. thetest compound in the absence of SB 203,186 in one tissue was comparedwith the test compound in the presence of SB 203,186 in a separatetissue. It was not possible to Carry out a curve 3 i.e. 5-HT curve 1,followed by the test compound curve 2 (−SB 203,186), followed by thetest compound curve 3 (+SB 203,186).

Agonist-Induced cAMP Elevation in Human 5-HT_(4(d)) transfected HEK293cells

Human 5-HT_(4(d)) transfected HEK293 cells were established in-house.The cells were grown at 37° C. and 5% CO₂ in DMEM supplemented with 10%FCS, 20 mM HEPES (pH 7.4), 200 μg/ml hygromycin B (Gibco), 100 units/mlpenicillin and 100 μg/ml streptomycin.

The cells were grown to 60-80% confluence. On the previous day beforetreatment with compounds dialyzed FCS (Gibco) was substituted for normaland the cells were incubated overnight.

Compounds were prepared in 96-well plates (12.5 μl/well). The cells wereharvested with PBS/1 mM EDTA, centrifuged and washed with PBS. At thebeginning of the assay, cell pellet was resuspended in DMEM supplementedwith 20 mM HEPES, 10 μM pargyline (Sigma) and 1 mM3-isobutyl-1-methylxanthine (Sigma) at the concentration of 1.6×10⁵cells/ml and left for 15 minutes at room temperature. The reaction wasinitiated by addition of the cells into plates (12.5 μl/well). Afterincubation for 15 minutes at room temperature, 1% Triton X-100 was addedto stop the reaction (25 μl/well) and the plates were left for 30minutes at room temperature. Homogenous time-resolved fluorescence-basedcAMP (Schering) detection was made according to the manufacturer'sinstruction. ARVOsx multilabel counter (Wallac) was used to measure HTRF(excitation 320 nm, emission 665 nm/620 nm, delay time 50 μs, windowtime 400 μs).

Data was analyzed based on the ratio of fluorescence intensity of eachwell at 620 nm and 665 nm followed by cAMP quantification using cAMPstandard curve. Enhancement of cAMP production elicited by each compoundwas normalized to the amount of cAMP produced by 1000 nM serotonin(Sigma).

All compounds of Examples showed 5HT₄ receptor agonistic activity.

Human Dofetilide Binding

Human HERG transfected HEK293S cells were prepared and grown in-house.The collected cells were suspended in 50 mM Tris-HCl (pH 7.4 at 4° C.)and homogenized using a hand held Polytron PT 1200 disruptor set at fullpower for 20 sec on ice. The homogenates were centrifuged at 48,000×g at4° C. for 20 min. The pellets were then resuspended, homogenized, andcentrifuged once more in the same manner. The final pellets wereresuspended in an appropriate volume of 50 mM Tris-HCl, 10 mM KCl, 1 mMMgCl₂ (pH 7.4 at 4° C.), homogenized, aliquoted and stored at −80° C.until use. An aliquot of membrane fractions was used for proteinconcentration determination using BCA protein assay kit (PIERCE) andARVOsx plate reader (Wallac).

Binding assays were conducted in a total volume of 200 μl in 96-wellplates. Twenty μl of test compounds were incubated with 20 μl of[³H]-dofetilide (Amersham, final 5 nM) and 160 μl of membrane homogenate(25 μg protein) for 60 minutes at room temperature. Nonspecific bindingwas determined by 10 μM dofetilide at the final concentration.Incubation was terminated by rapid vacuum filtration over 0.5% presoakedGF/B Betaplate filter using Skatron cell harvester with 50 mM Tris-HCl,10 mM KCl, 1 mM MgCl₂, pH 7.4 at 4° C. The filters were dried, put intosample bags and filled with Betaplate Scint. Radioactivity bound tofilter was counted with Wallac Betaplate counter.

I_(HERG) Assay

HEK 293 cells which stably express the HERG potassium channel were usedfor electrophysiological study. The methodology for stable transfectionof this channel in HEK cells can be found elsewhere (Z. Zhou et al.,1998, Biophysical journal, 74, pp 230-241). Before the day ofexperimentation, the cells were harvested from culture flasks and platedonto glass coverslips in a standard MEM medium with 10% FCS. The platedcells were stored in an incubator at 37° C. maintained in an atmosphereof 95% O₂/5% CO₂. Cells were studied between 15-28 hrs after harvest.

HERG currents were studied using standard patch clamp techniques in thewhole-cell mode. During the experiment the cells were superfused with astandard external solution of the following composition (mM); NaCl, 130;KCl, 4; CaCl₂, 2; MgCl₂, 1; Glucose, 10; HEPES, 5; pH 7.4 with NaOH.Whole-cell recordings was made using a patch clamp amplifier and patchpipettes which have a resistance of 1-3 MOhm when filled with thestandard internal solution of the following composition (mM); KCl, 130;MgATP, 5; MgCl₂, 1.0; HEPES, 10; EGTA 5, pH 7.2 with KOH. Only thosecells with access resistances below 15 MΩ and seal resistances >1 GΩ wasaccepted for further experimentation. Series resistance compensation wasapplied up to a maximum of 80%. No leak subtraction was done. However,acceptable access resistance depended on the size of the recordedcurrents and the level of series resistance compensation that can safelybe used. Following the achievement of whole cell configuration andsufficient for cell dialysis with pipette solution (>5 min), a standardvoltage protocol was applied to the cell to evoke membrane currents. Thevoltage protocol is as follows. The membrane was depolarized from aholding potential of −80 mV to +20 mV for 1000 ms. This was followed bya descending voltage ramp (rate 0.5 mV msec⁻¹) back to the holdingpotential. The voltage protocol was applied to a cell continuouslythroughout the experiment every 4 seconds (0.25 Hz). The amplitude ofthe peak current elicited around −40 mV during the ramp was measured.Once stable evoked current responses were obtained in the externalsolution, vehicle (0.5% DMSO in the standard external solution) wasapplied for 10-20 min by a peristalic pump. Provided there were minimalchanges in the amplitude of the evoked current response in the vehiclecontrol condition, the test compound of either 0.3, 1, 3, 10 μM wasapplied for a 10 min period. The 10 min period included the time whichsupplying solution was passing through the tube from solution reservoirto the recording chamber via the pump. Exposing time of cells to thecompound solution was more than 5 min after the drug concentration inthe chamber well reached the attempting concentration. Therereversibility. Finally, the cells was exposed to high dose of dofetilide(5 μM), a specific IKr blocker, to evaluate the insensitive endogenouscurrent.

All experiments were performed at room temperature (23±1° C.). Evokedmembrane currents were recorded on-line on a computer, filtered at 500-1KHz (Bessel −3dB) and sampled at 1-2 KHz using the patch clamp amplifierand a specific data analyzing software. Peak current amplitude, whichoccurred at around −40 mV, was measured off line on the computer.

The arithmetic mean of the ten values of amplitude was calculated undercontrol conditions and in the presence of drug. Percent decrease ofI_(N) in each experiment was obtained by the normalized current valueusing the following formula: I_(N)=(1−I_(D)/I_(C))×100, where I_(D) isthe mean current value in the presence of drug and I_(C) is the meancurrent value under control conditions. Separate experiments wereperformed for each drug concentration or time-matched control, andarithmetic mean in each experiment is defined as the result of thestudy.

Half-Life in Human Liver Microsomes (HLM)

Test compounds (1 μM) were incubated with 3.3 mM MgCl₂ and 0.78 mg/mLHLM (HL101) in 100 mM potassium phosphate buffer (pH 7.4) at 37° C. onthe 96-deep well plate. The reaction mixture was split into two groups,a non-P450 and a P450 group. NADPH was only added to the reactionmixture of the P450 group. An aliquot of samples of P450 group wascollected at 0, 10, 30, and 60 min time point, where 0 min time pointindicated the time when NADPH was added into the reaction mixture ofP450 group. An aliquot of samples of non-P450 group was collected at −10and 65 min time point. Collected aliquots were extracted withacetonitrile solution containing an internal standard. The precipitatedprotein was spun down in centrifuge (2000 rpm, 15 min). The compoundconcentration in supernatant was measured by LC/MS/MS system.

The half-life value was obtained by plotting the natural logarithm ofthe peak area ratio of compounds/internal standard versus time. Theslope of the line of best fit through the points yields the rate ofmetabolism (k). This was converted to a half-life value using followingequations:

Half-life=ln 2/k

Method of Gastric Emptying Model in Rats:

The effects of compounds on gastric emptying in rats were examined bythe modified method of D. A. Droppleman et al. (J. Pharmacol. Methods 4,227-230 (1980)). The test meal, non-fat caloric meal, was preparedaccording to the method of S. Ueki et al Arzneim.-Forsch./Drug Res. 49(II), 618-625 (1999)). IGS-SD rats (Male, 7 w, 230-270 g) were purchasedfrom Charles River Japan (Atsugi). These rats were used in theexperiments after one week acclimatization. In the experiments, ratswere fasted 15 hrs before the experiments but allowed free access towater. Forty-five minutes prior to the start of the experiment, waterwas removed from the cage to prevent rats from taking water. Fiveminutes before the test meal administration, test compounds, cisaprideor vehicle were dosed via an appropriate route to rats (n=8-10) in avolume of 0.1 ml per 100 g body weight. Cisapride (3 mg/kg) was used asa positive control for the experiment. Rats were given 3 ml of the testmeal by gavage and were returned to the cages. Thirty minutes after themeal administration, rats were culled by CO₂ exposure. Following amidline laparotomy, the stomach is ligated at the lower esophagealsphincter (LES) and pylorus. Then the stomach was removed and weighed(A). After the stomach was opened and rinsed with 0.9% saline, it wasblotted the face with the tissue to remove any excess liquid and weighedagain (B). After avoiding the rats that had eaten feces or givenartificial miss, gastric emptying rate for individual animals wascalculated by the formula:

GE rate (%)=(A−B)/weight of the test meal.

Gastric Motility in Conscious Dogs:

The surgical operation in dogs was performed by the modified method ofZ. Itoh et al. (Gastroenterol. Jpn., 12, 275-283 (1977)). The effects oftest compounds on gastric motility in dogs were examined by the modifiedmethod of N. Toshida et al. (J. Pharmacol. Exp/Ther., 257, 781-787(1991)).

An evaluation in the fasted state: Animals were chronically implantedwith a strain gauge force transducer on the gastric body, and fastedovernight prior to the experiment. The gastric motility was continuouslyrecorded by a telemetry system for 8 h after administration of thecompound. To quantitate the change in gastrointestinal motility, themotor index was determined as the area under the contraction curvesduring each 2 h period divided by the peak height of interdigestivemigrating contraction.

An evaluation in the postprandial state: Animals were chronicallyimplanted with a strain gauge force transducer on the gastric body, andfasted overnight prior to the experiment. Postprandial motility wasinduced by feeding with solid meal (100 grams), and the compound wasadministered 2 h later. The gastric motility was continuously recordedby a telemetry system for 8 h after administration of the compound. Themotor index was determined to quantitate the change in gastrointestinalmotility as the area under the contraction curves during each 1 h perioddivided by the area under the contraction curves for 1 h before thecompound administration.

The compounds of formula (I) of this invention can be administered viaeither the oral, parenteral or topical routes to mammals. In general,these compounds are most desirably administered to humans in dosesranging from 0.3 mg to 750 mg per day, preferably from 10 mg to 500 mgper day, although variations will necessarily occur depending upon theweight and condition of the subject being treated, the disease statebeing treated and the particular route of administration chosen.However, for example, a dosage level that is in the range of from 0.06mg to 2 mg per kg of body weight per day is most desirably employed fortreatment of inflammation.

The compounds of the present invention may be administered alone or incombination with pharmaceutically acceptable carriers or diluents byeither of the above routes previously indicated, and such administrationcan be carried out in single or multiple doses. More particularly, thenovel therapeutic agents of the invention can be administered in a widevariety of different dosage forms, i.e., they may be combined withvarious pharmaceutically acceptable inert carriers in the form oftablets, capsules, lozenges, troches, hard candies, powders, sprays,creams, salves, suppositories, jellies, gels, pastes, lotions,ointments, aqueous suspensions, injectable solutions, elixirs, syrups,and the like. Such carriers include solid diluents or fillers, sterileaqueous media and various non-toxic organic solvents, etc. Moreover,oralpharmaceutical compositions can be suitably sweetened and/orflavored. In general, the therapeutically-effective compounds of thisinvention are present in such dosage forms at concentration levelsranging 5% to 70% by weight, preferably 10% to 50% by weight.

For oral administration, tablets containing various excipients such asmicrocrystalline cellulose, sodium citrate, calcium carbonate,dipotassium phosphate and glycine may be employed along with variousdisintegrants such as starch and preferably corn, potato or tapiocastarch, alginic acid and certain complex silicates, together withgranulation binders like polyvinylpyrrolidone, sucrose, gelatin andacacia. Additionally, lubricating agents such as magnesium stearate,sodium lauryl sulfate and talc are often very useful for tablettingpurposes. Solid compositions of a similar type may also be employed asfillers in gelatin capsules; preferred materials in this connection alsoinclude lactose or milk sugar as well as high molecular weightpolyethylene glycols. When aqueous suspensions and/or elixirs aredesired for oral administration, the active ingredient may be combinedwith various sweetening or flavoring agents, coloring matter or dyes,and, if so desired, emulsifying and/or suspending agents as well,together with such diluents as water, ethanol, propylene glycol,glycerin and various like combinations thereof.

For parenteral administration, solutions of a compound of the presentinvention in either sesame or peanut oil or in aqueous propylene glycolmay be employed. The aqueous solutions should be suitably buffered(preferably pH>8) if necessary and the liquid diluent first renderedisotonic. These aqueous solutions are suitable for intravenous injectionpurposes. The oily solutions are suitable for intra-articular,intra-muscular and subcutaneous injection purposes. The preparation ofall these solutions under sterile conditions is readily accomplished bystandard pharmaceutical techniques well known to those skilled in theart. Additionally, it is also possible to administer the compounds ofthe present invention topically when treating inflammatory conditions ofthe skin and this may preferably be done by way of creams, jellies,gels, pastes, ointments and the like, in accordance with standardpharmaceutical practice.

Examples

The invention is illustrated in the following non-limiting examples inwhich, unless stated otherwise: all operations were carried out at roomor ambient temperature, that is, in the range of 18-25° C.; evaporationof solvent was carried out using a rotary evaporator under reducedpressure with a bath temperature of up to 60° C.; reactions weremonitored by thin layer chromatography (tlc) and reaction times aregiven for illustration only; melting points (m.p.) given are uncorrected(polymorphism may result in different melting points); the structure andpurity of all isolated compounds were assured by at least one of thefollowing techniques: tlc (Merck silica gel 60 F₂₅₄ precoated TLC platesor Merck NH₂ F_(254s) precoated HPTLC plates), mass spectrometry,nuclear magnetic resonance (NMR), infrared red absorption spectra (IR),microanalysis or powder X-ray diffraction(PXRD) pattern. Yields aregiven for illustrative purposes only. Flash column chromatography wascarried out using Merck silica gel 60 (230-400 mesh ASTM) or FujiSilysia Chromatorex® DU3050 (Amino Type, 30-50 μm). Low-resolution massspectral data (EI) were obtained on a Integrity (Waters) massspectrometer or a Automass 120 (JEOL) mass spectrometer. Low-resolutionmass spectral data (ESI) were obtained on a ZMD2 (Waters) massspectrometer or a Quattro II (Micromass) mass spectrometer. NMR data wasdetermined at 270 MHz (JEOL JNM-LA 270 spectrometer) or 300 MHz (JEOLJNM-LA300) using deuterated chloroform (99.8% D) or dimethylsulfoxide(99.9% D) as solvent unless indicated otherwise, relative totetramethylsilane (TMS) as internal standard in parts per million (ppm);conventional abbreviations used are: s=singlet, d=doublet, t=triplet,q=quartet, m=multiplet, br.=broad, etc. IR spectra were measured by aShimazu infrared spectrometer (IR-470). Optical rotations were measuredusing a JASCO DIP-370 Digital Polarimeter (Japan Spectroscopic CO,Ltd.). PXRD pattern was determined using a Rigaku RINT-TTR powder X-raydiffractometer fitted with an automatic sample changer, a 2 theta-thetagoniometer, beam divergence slits, a secondary monochromator and ascintillation counter. The sample was prepared for analysis by packingthe powder on to an aluminum sample holder. The specimen was rotated by60.00 rpm and scanned by 4°/min. Chemical symbols have their usualmeanings; b.p. (boiling point), m.p. (melting point), l (liter(s)), ml(milliliter(s)), g (gram(s)), mg(milligram(s)), mol (moles), mmol(millimoles), eq. (equivalent(s)).

Example 1N-({1-[(4-Hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

Step 1.tert-Butyl({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)carbamate

To a stirred solution of tert-butyl(piperidin-4-ylmethyl)carbamate (22.3g, 104 mmol) in methanol was added 1,6-dioxaspiro[2.5]octane (14.2 g,124 mmol, Satyamurthy, Nagichettiar et al., Phosphorus Sulfur, 1984, 19,113) at ambient temperature.

Then, the mixture was heated at 60° C. for 4 h. The volatile componentswere removed by evaporation and the resulting viscous oil wasprecipitated with a mixture of hexane and diethylether. The precipitatewas collected by filtration and recrystallized with a mixture of hexaneand 2-propanol to give title compound 14.2 g (42%) as a colorlesspowder.

MS (ESI) m/z: 329 (M+H⁺).

m.p.: 104° C.

¹H-NMR (CDCl₃) δ: 1.23-1.31 (2H, m), 1.44 (9H, s), 1.51-1.69 (8H, m),2.27-2.38 (4H, m), 2.83-2.88 (2H, m), 3.00 (2H, t, J=6.2 Hz), 3.70-3.85(4H, m).

Anal. Calcd. for C₁₇H₃₂N₂O₄: C, 62.17; H, 9.82; N, 8.53. Found: C,62.07; H, 9.92; N, 8.58.

Step 2.4-{[4-(Aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol

To a solution oftert-butyl({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)carbamate(50.28 g, 153 mmol) in methanol was added 4N HCl in dioxane (200 mL, 800mmol) at room temperature. After 4 h, the volatile materials wereremoved by evaporation. The resulting amorphous was precipitated withdiethyl ether/methanol (5:1). The precipitate was collected and added tothe ice cooled 6N NaOH aq. (200 mL) gradually. The mixture was extractedwith dichloromethane/methanol (10:1) for 4 times. The combined organicphase was washed with brine, dried over MgSO₄ and concentrated to give24.90 g (99%) of the title compound as pale brown amorphous.

MS (ESI) m/z: 229 (M+H⁺).

¹H-NMR (CDCl₃) δ: 1.19-1.28 (2H, m), 1.44-1.63 (8H, m), 1.65-1.71 (2H,m), 2.32 (2H, s), 2.35 (2H, t, J=11.0 Hz), 2.57 (2H, d, J=5.7 Hz),2.85-2.90 (2H, m), 3.70-3.81 (4H, m).

Step 3.N-({1-[(4-Hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

To a stirred mixture of 1-isopropyl-1,3-dihydro-2H-benzimidazol-2-one(J. Med. Chem. 1999, 42, 2870-2880) (23.0 g, 130 mmol) and triethylamine(54.6 mL, 392 mmol) in tetrahydrofuran (300 mL) was added triphosgen(38.8 g, 130 mmol) in tetrahydrofuran (200 mL) gradually at roomtemperature. Then, the mixture was heated at 80° C. for 4 h. Aftercooling, a solution of4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol (step2 of Example 1) (24.9 g, 109 mmol) and triethylamine (45 mL, 109 mmol)in tetrahydrofuran (500 mL) was added to the mixture. Then, the mixturewas heated at 80° C. for 6 h. After cooling, sat. NaHCO₃ aq was added tothe mixture. The mixture was extracted with ethyl acetate (500 mL×4).The extracts were washed with brine, dried over MgSO₄ and concentrated.The residue was chromatographed on a column of aminopropyl-silica geleluting with hexane/ethyl acetate (3:1) to give 31.3 g (67%) of thetitle compound as a white solid.

¹H NMR (DMSO-d₆) δ 8.80 (1H, br t, J=6.0 Hz), 8.06 (1H, m), 7.41 (1H,m), 7.19 (1H, dt, J=1.5, 7.7 Hz), 7.12 (1H, dt, J=1.3, 7.7 Hz), 4.64(1H, septet, J=7.0 Hz), 4.08 (1H, br s), 3.68-3.44 (4H, m), 3.19 (2H, t,J=6.0 Hz), 2.89 (2H, m), 2.20 (2H, br s), 2.09 (2H, m), 1.68-1.10 (9H,m), 1.47 (6H, d, J=7.0 Hz).

MS (ESI) m/z: 431 (M+H⁺).

Anal. Calcd. for C23H34N4O4: C, 64.16; H, 7.96; N, 13.01. Found: C,64.13; H, 7.97; N, 12.99.

Step 4.N-({1-[(4-Hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

To a stirred solution ofN-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide(27.0 g, 62 mmol) in methanol (150 mL) was added 10% HCl-methanol (100mL) at ambient temperature. After 30 min, the volatile materials wereremoved by evaporation. The resulting amorphous was precipitated byethanol/diethylether. The precipitate was recrystallized fromethanol/diethylether (1:1) to give 26.5 g (90%) of title compound as acolorless powder.

¹H-NMR (DMSO-d₆) δ: 1.49 (6H, d, J=6.9 Hz), 1.50-1.70 (4H, m), 1.76-1.91(5H, m), 3.00-3.12 (3H, m), 3.15-3:45 (3H, m), 3.60-3.70 (6H, m),4.61-4.69 (1H, m), 5.46-5.49 (1H, m), 7.13 (1H, t, J=7.8 Hz) 7.20 (1H,t, J=7.8 Hz), 7.42 (1H, d, J=7.9 Hz), 8.07 (1H, d, J=8.0 Hz), 8.86 (1H,m), 9.61-9.81 (1H, m)

MS (ESI) m/z: 431 (M+H⁺).

Anal. Calcd. for C23H35N4O4Cl: C, 59.15; H, 7.55; N, 2.00. Found: C,58.81; H, 7.57; N, 11.85.

Alternative route to synthesize4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol isdescribed below.

Step 1.Benzyl({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)carbamate

A mixture of benzyl(piperidin-4-ylmethyl)carbamate (7.77 g, 31.3 mmol ,Bose, D. Subhas et al., Tetrahedron Lett., 1990, 31, 6903) and1,6-dioxaspiro[2.5]octane (4.29 g, 37.6 mmol, Satyamurthy, Nagichettiaret al., Phosphorus Sulfur, 1984, 19, 113) in methanol (93 mL) wasstirred at room temperature for 20 h. Then the mixture was refluxed for8 h. After cooling to room temperature, the solvent was removed invacuo. The residue was chromatographed on a column of silica gel elutingwith methanol/dichloromethane (1:20) to give 5.60 g (49%) of the titlecompound as a colorless oil.

Step 2.4-{[4-(Aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol

A mixture ofbenzyl({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)carbamate(5.60 g, 15.5 mmol, step 1) and palladium on activated carbon (10 wt. %,1.20 g) in methanol (250 mL) was hydrogenated at room temperature for 20h. Then, the mixture was filtered through a pad of Celite, and thefiltrate was concentrated in vacuo to give 3.30 g (94%) of the titlecompound as slightly yellow oil.

Following is an another route to synthesize4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol.

Step 1.1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidine-4-carboxamide

The mixture of trimethylsulfoxonium iodide (0.791 g, 3.52 mmol) and2N—NaOH aq (1.76 mL, 3.52 mmol) in acetonitrile (1.62 mL) was stirred at50° C. for 30 min. Then to the mixture was addedtetrahydro-4H-pyran-4-one (0.324 g, 3.20 mmol) and the resulting mixturewas stirred at 50° C. for 3 h. Sat. NaCl aq. (10 mL) was added to thereaction mixture at room temperature and organic layer was extractedwith CH₂Cl₂ (20 mL), dried over Na₂SO₄, filtered and concentrated. Afterremoval of the solvent, MeOH (1.62 mL) and isonipecotamide (0.381 g,2.88 mmol) were added to the residue, the mixture was stirred at 75° C.for 14 h under N₂. The reaction mixture was concentrated and the residuewas recrystallized from MeOH-acetonitrile to give 0.484 g (2.00 mmol) oftitle compound as a white solid.

¹H-NMR (300 MHz, DMSO-d6) δ 7.19 (br s, 1H), 6.69 (br s, 1H), 4.10 (s,1H), 3.70-3.50 (m, 4H), 2.95-2.85 (m, 2H), 2.20 (s, 2H), 2.15-1.85 (m,3H), 1.65-1.50 (m, 6H), 1.40-1.25 (m, 2H).

Step 2.4-{[4-(Aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-oltosylate

To a stirred suspension of NaBH₄ (0.505 g, 13.2 g) in triethylene glycoldimethyl ether (12.8 mL) was added the solution1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidine-4-carboxamide(0.640 g, 2.64 mmol) and AcOH (0.765 mL, 13.2 mmol) in triethyleneglycol dimethyl ether (3.2 mL) dropwise at 80° C. under N₂. The reactionmixture was quenched with 2N—HCl aq until pH value was <3, then theresulting mixture was stirred at room temperature for 1 h. To themixture CH₂Cl₂ (30 mL) and 2N—NaOH aq. was added until pH value ofaqueous layer was >10. Organic layer was extracted with CH₂Cl₂ for threetimes, and the combined organic layer was dried over Na₂SO₄, filteredand concentrated.

To the residual solution (title compound in triethylene glycol dimethylether) the solution of p-toluenesulfonic acid monohydrate (0.408 g, 2.11mmol) in MeOH (1.28 mL) was added at 60° C., then the mixture was cooledto room temperature. Appeared solids were collected by suction and washwith hexane to give title compound (0.340 g, 0.849 mmol) as a whitesolid

¹H-NMR (300 MHz, DMSO-d6) δ 7.61 (br s, 2H), 7.55-7.40 (m, 2H),7.15-7.05 (m, 2H), 4.11 (br s, 1H), 3.70-3.45 (m, 4H), 2.95-2.85 (m,2H), 2.68 (d, J=7.0, 2H), 2.29 (s, 3H), 2.22 (s, 2H), 2.07 (t, J=11.0,2H), 1.65-1.45 (m, 4H), 1.55-1.35 (m, 1H), 1.40-1.25 (m, 2H), 1.30-1.10(m, 2H).

Example 2N-({1-[(4-Hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehemiedisylate

To a stirred solution ofN-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide1.51 g (3.51 mmol) in ethyl acetate (10 mL) and methanol (10 mL) wasadded a solution of 1,2-ethanedisulfonic acid dihydrate 397 mg (1.75mmol) in methanol (5.0 mL) and the resulting suspension was stirred for5 h at room temperature. The mixture was filtered and the first crop wasdried under vacuum for 5 h at 100° C. to give 1.78 g of crude product.1.61 g of the crude product was dissolved in methanol (20 mL) and ethylacetate (20 mL) was added to the solution. The resulting suspension wasstirred for 2 h at room temperature. The mixture was filtered and thecrop was dried under vacuum for 4 h at 100° C. to give the titledcompound 1.13 g (61%) as colorless crystals.

MS (ESI) m/z: 431 (M+H)⁺.

m.p.: 233° C.

IR (KBr) v: 2866, 1738, 1683, 1558, 1373, 1217, 1028, 756 cm⁻¹.

¹H NMR (DMSO-d₆) δ 8.96 (0.25H, br s), 8.85 (1H, br t, J=6.0 Hz), 8.61(0.75H, br s), 8.06 (1H, m), 7.43 (1H, m), 7.21 (1H, dt, J=1.3, 7.7 Hz),7.13 (1H, dt, J=1.2, 7.7 Hz), 5.26 (1H, br s), 4.65 (1H, septet, J=7.0Hz), 3.74-2.92 (12H, m), 2.64 (2H, s), 2.00-1.35 (9H, m), 1.47 (6H,d,J=7.0 Hz).

Anal. calcd. for C₂₃H₃₄N₄O₄.0.5C₂H₆O₆S₂: C, 54.84; H, 7.09; N, 10.66; S,6.10. Found: C, 54.50; H, 7.24; N, 10.60; S, 6.08.

PXRD pattern angle (2-Theta°): 10.2, 11.9, 16.3, 17.3, 17.6, 21.8, 24.2.

Example 33-Isopropyl-N-{[1-(2-morpholin-4-yl-2-oxoethyl)piperidin-4-yl]methyl}-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidemonooxalate

The tilted compound was prepared with the similar method shown in theStep 3 of Preparation 1 by using 4-(chloroacetyl)morpholine (B. G.Hazra; V. S. Pore; S. P. Maybhate, Org. Prep. Proced. Int., 1989, 21,355-8).

MS (ESI) m/z: 440 (M+H)⁺.

m.p.: 194.2° C.

IR (KBr) v: 3443, 2934, 1765, 1728, 1686, 1659, 1612, 1551 cm⁻¹.

¹H-NMR (CDCl₃) (free base) δ: 9.00-8.88 (1H, m) 8.30-8.22 (1H, m),7.23-7.12 (3H, m), 4.78-4.62 (1H, m), 3.66 (4H, s), 3.70-3.58 (4H, m),3.32 (2H, t, J=6.3 Hz), 3.15 (2H, s), 2.94-2.84 (2H, m), 2.14-2.01 (2H,m), 1.86-1.23 (5H, m), 1.56 (6H, d, J=7.0 Hz).

¹H-NMR (DMSO-d₆) (salt form) δ: 8.92-8.80 (1H, m) 8.07 (1H, d, J=7.7Hz), 7.45 (1H, d, J=7.5 Hz), 7.26-7.06 (2H, m), 4.76-4.56 (1H, m),4.10-2.60 (18H, m), 1.90-1.40 (3H, m), 1.49 (6H, d, J=6.9 Hz).

Anal. Calcd. for C₂₅H₃₅N₅O₈: C, 56.27; H, 6.61; N, 13.13. Found: C,56.25; H, 6.82; N, 12.98.

Example 43-Isopropyl-N-{[1-(3-morpholin-4-yl-3-oxopropyl)piperidin-4-yl]methyl}-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidemonooxalate

A mixture of3-isopropyl-2-oxo-N-(piperidin-4-ylmethyl)-2,3-dihydro-1H-benzimidazole-1-carboxamide(150 mg, 0.474 mmol) and 4-(3-chloro-propanoyl)-morpholine (G. Mattalia;C. Serafini; U. Bucciarelli, Farmaco, Ed. Sci., 1976, 31, 457-67) (300mg, 1.185 mmol) in 4.7 ml N,N-dimethylformamide was added triethylamine(0.23 ml, 1.659 mmol) and sodium iodide (178 ml, 1.185 mmol). Thereaction mixture was stirred at 90° C. for 6 days. The reaction mixturewas then concentrated by evaporation. The residue was diluted aqueousNaHCO₃ 10 ml, extracted with dichloromethane 30 ml for three times. Thecombined extract was dried over MgSO₄ and concentrated. Preparative TLC(elutent: CH₂Cl₂/methanol=10/1) afforded a brown amorphous oil 130 mg(60%). The amorphous (130 mg) was dissolved in 3 ml methanol andacidified with a solution of 24 mg oxalic acid in 2 ml MeOH. The mixturewas concentrated. Crystallization of the resulting residue withAcOEt-EtOH afforded a white amorphous 107 mg as the titled compound.

MS (ESI) m/z: 458 (M+H)⁺.

IR (KBr) v: 3443, 2941, 1732, 1697, 1686, 1647, 1638, 1558 cm⁻¹.

¹H-NMR (CDCl₃) (free base) δ: 9.06-8.94 (1H, br) 8.24-8.19 (1H, m),7.26-7.10 (3H, m), 4.76-4.64 (1H, m), 3.75-2.80 (10H, m), 2.60-1.30(13H, m), 1.56 (6H, d, J=7.0 Hz).

¹H-NMR (CDCl₃) (salt form) δ: 9.10-9.00 (1H, m) 8.27-8.17 (1H, m),7.33-7.12 (3H, m), 4.87-4.62 (1H, m), 3.78-2.65 (16H, m), 2.20-1.60 (7H,m), 1.56 (6H, d, J=6.9 Hz).

Anal. Calcd. for C₂₆H₃₇N₅O₈.0.9C₂H₂O₄.1.3H₂O: C, 51.21; H, 6.40; N,10.74. Found: C, 50.90; H, 6.26; N, 11.13.

Example 53-Isopropyl-N-{[1-(4-morpholin-4-yl-4-oxobutyl)piperidin-4-yl]methyl}-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidemonooxalate

The tilted compound was prepared with the similar method shown in theexample 4 by using 4-(4-chloro-butyryl)-morpholine (Schlesinger; Prill;B. G. Hazra; J. Amer. Chem. Soc., 1956, 78, 6123-6124).

MS (ESI) m/z: 472 (M+H)⁺.

IR (KBr) v: 3443, 1728, 1686, 1647-1616, 1551 cm⁻¹.

¹H-NMR (CDCl₃) (free base) δ: 9.02-8.88 (1H, m) 8.31-8.20 (1H, m),7.22-7.04 (3H, m), 4.80-4.60 (1H, m), 3.66-3.56 (8H, m), 3.40-3.22 (2H,m), 3.00-2.88 (2H, m), 2.50-2.30 (6H, m), 2.00-1.20 (7H, m), 1.57 (6H,d, J=7.1 Hz).

¹H-NMR (DMSO-d₆) (salt form) δ: 8.93-8.79 (1H, m) 8.07 (1H, d, J=7.5Hz), 7.44 (1H, d, J=7.5 Hz), 7.27-7.08 (2H, m), 4.75-4.58 (1H, m),4.47-2.30 (18H, m), 1.90-0.90 (7H, m), 1.49 (6H, d, J=6.9 Hz).

Anal. Calcd. for C₂₇H₃₉N₅O₈: C, 57.74; H, 7.00; N, 12.47. Found: C,57.52; H, 7.03; N, 12.32.

Example 6N-({1-[(trans-1,4-Dihydroxyhexyl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

Step 1. tert-Butyl(1-oxaspiro[2.5]oct-6-yloxy)diphenylsilane

To a stirred suspension of sodium hydride (60% in mineral oil, 441 mg,11.0 mmol) in DMSO (7 ml) was added trimethylsulfoxonium iodide (2.53 g,11.5 mmol) at room temperature, and the mixture was stirred at roomtemperature for 30 min. To this mixture was added a solution of4-{[tert-butyl(diphenyl)silyl]oxy}cyclohexanone (Okamura, William H. etal., J. Org. Chem., 1993, 58, 600-610, 3.53 g, 10.0 mmol) in DMSO (35ml) dropwise at room temperature, the mixture was stirred at roomtemperature for 2 h. Then the mixture was diluted with water (600 ml),and extracted with diethylether (200 ml×4). The combined organic layerwas dried over magnesium sulfate, and concentrated in vacuo. The residuewas chromatographed on a column of silica gel eluting withn-hexane/ethyl acetate (1:10), and then purified with PTLC eluting withn-hexane/ethyl acetate (1:15) to give 459 mg (13%, trans) and 390 mg(11%, cis) of the title compound as colorless oil respectively.

(trans)

¹H-NMR (CDCl₃) δ: 7.70-7.66 (4H, m), 7.46-7.35 (6H, m), 4.03-3.97 (1H,m), 2.63 (2H, s), 2.07-1.63 (8H, m), 1.08 (9H, s).

(cis)

¹H-NMR (CDCl₃) δ: 7.70-7.65 (4H, m), 7.46-7.35 (6H, m), 3.97-3.83 (1H,m), 2.58 (2H, s), 1.83-1.37 (8H, m), 1.07 (9H, s).

Step 2.N-{[1-({trans-4-[tert-Butyl(diphenyl)silyl]oxy-1-hydroxycyclohexyl}methyl)piperidin-4-yl]methyl}-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

A mixture oftert-butyl[(3R,6R)-1-oxaspiro[2.5]oct-6-yloxy]diphenylsilane (Step 1,trans-isomer, 283.0 mg, 0.772 mmol) and3-isopropyl-2-oxo-N-(piperidin-4-ylmethyl)-2,3-dihydro-1H-benzimidazole-1-carboxamide(Preparation 1; step 2, 2.48 g, 0.0194 mol) in MeOH (4 ml) was heated at50° C. with stirring for 2 days. After cooling, the reaction mixture wasevaporated to remove the solvent, and residue was chromatographed on acolumn of silica gel eluting with ethyl acetate/n-hexane (1:10) thenmethanol/dichloromethane (1:20) to give 308.1 mg (58%) of the titlecompound as a colorless syrup.

MS (ESI) m/z: 683 (M+H)⁺.

1H-NMR (CDCl₃) δ: 8.93 (1H, m), 8.32-8.23 (1H, m), 7.72-7.60 (4H, m),7.46-7.32 (6H, m), 7.22-7.10 (3H, m), 4.80-4.62 (1H, m), 3.96 (1H, m),3.31 (2H, t, J=6.26 Hz), 2.92 (2H, d, J=10.88 Hz), 2.45-2.29 (4H, m),1.85-1.65 (6H, m), 1.65-1.43 (9H, m, including 6H, d, J=7.09 Hz at 1.56ppm), 1.43-1.25 (4H, m), 1.06 (9H, s).

Step 3.N-({1-[(trans-1,4-Dihydroxycyclohexyl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

A mixture of tert-butylN-{[1-({trans-4-[tert-butyl(diphenyl)silyl]oxy-1-hydroxycyclohexyl}methyl)piperidin-4-yl]methyl}-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide(234 mg, 0.343 mol) and HCl solution of MeOH (50 ml) was stirred at roomtemperature for 4 h. Then the solvent was removed in vacuo. The residuewas basified with saturated aqueous NaHCO₃ (30 ml), extracted withCH₂Cl₂ (30 ml×3 times) and the combined organic layer was dried overNa₂SO₄. Removal of the solvent gave a residue, which was chromatographedon a column of NH-silica gel eluting with ethyl acetate/n-hexane(1:1-2:1) to give 140.1 mg (92%) of the title compound as a colorlesssyrup.

MS (ESI) m/z: 445 (M+H)⁺.

¹H NMR (CDCl₃) δ: 8.93 (1H, br t, J=5.87 Hz), 8.32-8.20 (1H, m),7.25-7.03 (3H, m), 4.80-4.62 (1H, m), 3.94 (1H, m), 3.31 (2H, t, J=6.10Hz), 2.89 (2H, br d, J=11.53 Hz), 2.36 (2H, s), 2.34 (2H, t, J=11.86Hz), 2.00-1.85 (2H, 1.82-1.25 (18H, m, including 6H, d, J=7.09 Hz at1.56 ppm).

140.1 mg of this syrup was dissolved in HCl solution in MeOH (4 ml),concentrated, and dried in vacuo at 50° C. for 5 h to give 139.2 mg oftitle compound as a yellow amorphous solid.

MS (ESI) m/z: 445 (M+H)⁺.

¹H NMR (DMSO-d6) δ: 9.35-8.75 (1H, m), 8.86 (1H, t, J=6.59 Hz), 8.07(1H, d, J=7.74 Hz), 7.44 (1H, d, J=7.58 Hz), 7.22 (1H, dt, J=1.15 Hz,7.42 Hz), 7.14 (1H, dt, J=1.32 Hz, 7.74 Hz), 5.04 (1H, br s), 4.75-4.45(1H, m), 3.70 (1H, br s), 3.59 (2H, d, J=11.70 Hz), 3.50-2.90 (8H, m),1.90-1.57 (8H, m), 1.57-1.30 (10H, m, including 6H, d, J=6.92 Hz at 1.49ppm)

IR(KBr): 3285, 2936, 2677, 1728, 1686, 1611, 1549, 1481, 1375, 1298,1204, 1157, 1101, 1018, 762 cm⁻¹

Anal. Calcd for C₂₄H₃₆N₄O₄—HCl-2H₂O: C, 57.76; H, 7.88; N, 11.23. Found:C, 57.54; H, 7.90; N, 11.21.

PXRD pattern angle (2-Theta°): 8.3, 14.5, 17.7, 18.3, 19.1, 26.4, 27.5.

Example 7N-({1-[(cis-1,4-Dihydroxyhexyl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

Step 1.N-{[1-({cis-4-[tert-Butyl(diphenyl)silyl]oxy-1-hydroxycyclohexyl}methyl)piperidin-4-yl]methyl}-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

The title compound was prepared according to the procedure described ofStep 2 in the Example 6 usingtert-butyl[(3S,6S)-1-oxaspiro[2.5]oct-6-yloxy]diphenylsilane (Example 6,Step 1, cis-isomer, 311.0 mg, 0.848 mmol) instead oftert-butyl[(3R,6R)-1-oxaspiro[2.5]oct-6-yloxy]diphenylsilane.

MS (ESI) m/z: 683 (M+H)⁺.

¹H-NMR (CDCl₃) δ: 8.91 (1H, t, J=5.87 Hz), 8.30-8.22 (1H, m), 7.72-7.63(4H, m), 7.45-7.30 (6H, m), 7.20-7.10 (3H, m), 4.80-4.63 (1H, m), 3.59(1H, m), 3.29 (2H, t, J=6.24 Hz), 2.83 (2H, d, J=11.74 Hz), 2.26 (2H, t,J=11.55 Hz), 2.18 (2H, s), 1.85-1.65 (4H, m), 1.65-1.50 (11H, m,including 6H, d, J=7.15 Hz at 1.56 ppm), 1.40-1.30 (2H, m), 1.15-1.00(11H, m, including 9H, s, 1.05 ppm).

Step 2.N-({1-[(cis-1,4-Dihydroxycyclohexyl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

The title compound was prepared according to the procedure described ofStep 3 in the Example 6 usingN-{[1-({cis-4-[tert-butyl(diphenyl)silyl]oxy-1-hydroxycyclohexyl}methyl}piperidin-4-yl]methyl}-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide(295.0 mg, 0.432 mmol) instead ofN-{[1-({trans-4-[diphenyl(trimethylsilyl)methoxy]-1-hydroxycyclohexyl}methyl)piperidin-4-yl]methyl}-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide.

MS (ESI) m/z: 445 (M+H)⁺.

¹H NMR (CDCl3) δ: 8.93 (1H, br t, J=5.60 Hz), 8.31-8.22 (1H, m),7.25-7.10 (3H, m), 4.80-4.62 (1H, m), 3.63-3.49 (1H, m), 3.31 (2H, t,J=6.10 Hz), 2.89 (2H, br d, J=11.54 Hz), 2.33 (2H, dt, J=1.81 Hz, 11.70Hz), 1.85-1.60 (16H, m, including 6H, d, J=7.09 Hz at 1.57 ppm),1.45-1.18 (4H, m).

165.7 mg of this syrup was dissolved in HCl solution in MeOH (4 ml),concentrated, and dried in vacuo at 50° C. for 5 h to give 164.7 mg oftitle compound as a yellow amorphous solid.

MS (ESI) m/z: 445 (M+H)⁺.

¹H NMR (DMSO-d6) δ: 9.30-8.90 (1H, m), 8.86 (1H, t, J=5.93 Hz), 8.07(1H, d, J=7.58 Hz), 7.44 (1H, d, J=7.58 Hz), 7.22 (1H, dt, J=1.48 Hz,7.75 Hz), 7.15 (1H, dt, J=1.15 Hz, 7.74 Hz), 4.75-4.58 (1H, m),3.70-2.90 (11H, m), 1.90-1.67 (6H, m), 1.67-1.20 (12H, m, including 6H,d, J=6.92 Hz at 1.49 ppm).

IR(KBr): 3294, 2936, 2673, 1728, 1686, 1611, 1545, 1479, 1375, 1298,1203, 1158, 1134, 1101, 1051, 762 cm⁻¹

Anal. Calcd for C₂₄H₃₆N₄O₄—HCl-5H₂O: C, 54.79; H, 8.05; N, 10.65. Found:C, 54.75; H, 7.88; N, 10.56.

Example 86-Fluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

Step 1.6-Fluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

The title compound was prepared according to the procedure described inStep 3 of Example 1 from5-fluoro-1-isopropyl-1,3-dihydro-2H-benzimidazol-2-one (I. Tapia et al.,J. Med. Chem., 1999, 42, 2880.) and4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol (step2 of Example 1).

MS (ESI) m/z: 449 (M+H⁺).

¹H-NMR (CDCl₃) δ: 1.12-1.70 (8H, m), 1.55 (6H, d, J=7.0 Hz), 1.74 (2H,brd, 12.8 Hz), 2.31 (2H, s), 2.35 (2H, brt, J=11.9 Hz), 2.88 (2H, brd,J=11.7 Hz), 3.30 (2H, t, J=6.2 Hz), 3.70-3.85 (4H, m), 4.62-4.75 (1H,m), 6.90 (1H, td, J=9.0, 2.4 Hz), 7.02-7.07 (1H, m), 8.05 (1H, dd,J=9.5, 2.6 Hz), 8.85-8.92 (1H, m).

Step 2.6-Fluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

The title compound was prepared according to the procedure described inStep 4 of Example 1 from6-fluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide(step 1 of Example 8).

MS (ESI) m/z: 449 (M+H⁺).

¹H-NMR (DMSO-d₆) δ: 1.46 (6H, d, J=6.9 Hz), 1.55-1.65 (4H, m), 1.70-1.91(4H, m), 2.90-3.28 (8H, m), 3.50-3.67 (6H, m), 4.56-4.69 (1H, m),5.30-5.37 (1H, m), 5.76 (1H, s), 7.08 (1H, td, J=9.0, 2.4 Hz), 7.44-7.49(1H, m), 7.85 (1H, dd, J=9.5, 2.5 Hz), 8.81-8.85 (1H, m).

Anal. Calcd. for C₂₃H₃₄FN₄O₄Cl: C, 56.96; H, 7.07; N, 11.55. Found: C,57.00; H, 7.20; N, 11.43.

PXRD pattern angle (2-Theta°): 10.0, 14.6, 16.2, 18.5, 23.2, 25.3, 27.3.

Example 95-fluoro-N-({1-[(1(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

Step 1. (5-fluoro-2-nitrophenyl)isopropylamine

To a stirred mixture of 2,4-difluoro-1-nitrobenzene (4.77 g, 30 mmol)and K2CO3 (4.14 g, 30 mmol) in THF (30 mL) was added isopropyl amine(1.77 g, 30 mmol) in THF (10 mL) at 0° C. After being stirred for 13 h,the insoluble materials were removed by pad of Celite and the filtratewas concentrated under reduced pressure to give title compound (5.25 g,88%) as a pale yellow oil.

MS (ESI) m/z: 405 (M+H⁺).

¹H NMR (CDCl₃): δ 8.21 (1H, dd, J=9.3, 6.0 Hz), 6.48 (1H, dd, J=11.7,2.6 Hz), 6.39-6.29 (1H, m), 3.81-3.66 (1H, m), 1.33 (6H, d, J=6.4 Hz)

Step 2. 6-fluoro-1-isopropyl-1,3-dihydro-2H-benzimidazol-2-one

A mixture of (5-fluoro-2-nitrophenyl)isopropylamine (Step 1 of Example9, 5.85 g, 30 mmol) and 10% Pd—C (600 mg) in MeOH was stirred underatmosphere of hydrogen gas at at room temperature for 12 h. The catalystwas filtered off on a pad of Celite, and the filtrate was evaporatedunder reduced pressure. To the residue was added1,1′-carbonyldiimidazole (4.5 g, 28 mmol) and THF (100 mL) and thenstirred at 100° C. for 10 h. After cooling, the volatile materials wereremoved under reduced pressure and the residue was partitioned betweenethylacetate and H₂O. After extraction with ethylacetate (3 times), thecombined organic phase was washed with brine, dried over MgSO₄ andconcentrated. The residue was chromatographed on a column of silica geleluting with hexane/ethyl acetate (2:1) to give 3.47 g (60%) of thetitle compound as a white solid.

MS (ESI) m/z: 195 (M+H⁺), 193 (M−H⁺).

¹H NMR (CDCl₃): δ 7.06-6.99 (1H, m), 6.90 (1H, dd, J=9.2, 2.4 Hz),6.82-6.72 (1H, m), 4.83-4.62 (1H, m), 1.54 (6H, d, J=7.1 Hz)

Step 3.5-fluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

To a stirred mixture of6-fluoro-1-isopropyl-1,3-dihydro-2H-benzimidazol-2-one (Step 2 ofExample 9, 0.58 g, 3 mmol) and p-nitrophenylchloroformate (0.66 g, 3.3mmol) in dichloromethane (15 mL) was added triethylamine (1.25 mL, 9.0mmol) at room temperature. After being stirred for 2 h, a solution of4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol (Step2 of Example 1, 0.75 g, 3.3 mmol) in dichloromethane (15 mL) was addedto the mixture. After being stirred for 4 h, the mixture was dilutedwith ethyl acetate (100 mL). Then, the organic layer was washed with 0.5N NaOH aq. (10 mL) for 5 times and brine, dried over MgSO₄ andconcentrated. The residue was chromatographed on a column ofaminopropyl-silica gel eluting with hexane/ethyl acetate (3:1) to give0.97 g (79%) of the title compound as a white solid.

MS (ESI) m/z: 449 (M+H⁺).

¹H NMR (CDCl₃): δ 8.84-8.74 (1H, m), 8.21-8.11 (2H, m), 7.02-6.91 (2H,m), 4.68-4.56 (1H, m), 3.87-3.72 (4H, m), 3.34-3.25 (2H, m), 2.93-2.82(2H, m), 2.42-2.25 (4H, m), 1.79-1.68 (2H, m), 1.67-1.29 (13H, m).

Anal. calcd. for C₂₃H₃₃N₄O₄F: C, 61.59; H, 7.42; N, 12.49. Found: C,61.45; H, 7.33; N, 12.40.

Example 105,6-difluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

Step 1. 4,5-difluoro-N-isopropyl-2-nitroaniline

4,5-difluoro-2-nitroaniline (3.48 g, 20 mmol), 2,2-dimethoxypropane(11.9 mL, 100 mmol), and trifluoroacetic acid (1.6 mL, 21 mmol) weredissolved in toluene (40 mL) and stirred at room temperature for 1 h. Aboron-pyridine complex (2.12 mL, 21 mmol) was slowly added. The reactionmixture was stirred for 20 h. The solvent was evaporated in vacuo, andthe residue was taken up into water and extracted with dichloromethane.The organic extract was dried (Na₂SO₄) and concentrated in vacuo. Theresidue was chromatographed on a column of aminopropyl-silica geleluting with hexane/ethyl acetate (30:1) to give 2.42 g (56%) of thetitle compound as a bright orange solid.

¹H NMR (CDCl₃): δ 8.05 (1H, dd, J=10.8, 8.6 Hz), 6.61 (1H, dd, J=12.6,6.8 Hz), 3.77-3.62 (1H, m), 1.33 (6H, d, J=6.2 Hz).

Step 2. 5.6-difluoro-1-isopropyl-1,3-dihydro-2H-benzimidazol-2-one

The title compound was prepared according to the procedure described inStep 2 of Example 9 from 4,5-difluoro-N-isopropyl-2-nitroaniline (Step 1of Example 10).

MS (ESI) m/z: 213 (M+H⁺), 211 (M+H⁺).

¹H NMR (CDCl₃): δ 7.00-6.89 (2H, 4.76-4.57 (1H, 3.86-3.69 (4H, m), 3.31(2H, t, J=7.0 Hz), 2.95-2.82 (2H, m), 2.35 (2H, t, J=, 13.7 Hz), 2.31(2H, s), 1.67-1.25 (10H, m), 1.55 (6H, d, J=7.7 Hz).

Step 3.5,6-difluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

The title compound was prepared according to the procedure described inStep 3 of Example 9 from5,6-difluoro-1-isopropyl-1,3-dihydro-2H-benzimidazol-2-one (Step 2 ofExample 10) and4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol (Step2 of Example 1).

MS (ESI) m/z: 467 (M+H⁺).

¹H NMR (CDCl₃): δ 8.88-8.78 (1H, m), 8.25-8.15 (1H, m), 6.94-6.79 (2H,m), 4.73-4.57 (1H, m), 3.86-3.69 (4H, m), 3.31 (2H, t, J=7.0 Hz),2.95-2.82 (2H, m), 2.35 (2H, t, J=, 13.7 Hz), 2.31 (2H, s), 1.67-1.25(10H, m), 1.55 (6H, d, J=7.7 Hz).

Step 4.5.6-difluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

A mixture of5,6-difluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide(Step 3 of Example 10, 113 mg, 0.242 mmol) and 10% HCl-methanol (5 mL)was stirred for 1 h. Then, the volatile components were removed underreduced pressure and the residue was recrystallized from ethanol-diethylether to give 88 mg (72%) of the title compound as a colorless powder.

MS (ESI) m/z: 467 (M+H⁺).

¹H NMR (DMSO-d₆): δ8.82-8.71 (1H, m), 8.08-7.93 (1H, m), 7.78-7.67 (1H,m), 5.35-5.26 (1H, m), 4.69-4.52 (1H, m), 3.70-3.51 (6H, m), 3.41-2.91(7H, m), 1.94-1.53 (8H, m), 1.45 (6H, d, J=7.0 Hz).

Anal. calcd. for C₂₃H₃₃N₄O₄F₂Cl.1H₂O: C, 53.96; H, 6.69; N, 10.94.Found: C, 53.67; H, 6.64; N, 10.89.

Example 116-chloro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

Step 1.6-chloro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

The title compound was prepared according to the procedure described inStep 3 of Example 9 from5-chloro-1-isopropyl-1,3-dihydro-2H-benzimidazol-2-one (I. Tapia et al.,J. Med. Chem., 42, 2880 (1999)) and4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol (Step2 of Example 1).

MS (ESI) m/z: 465 (M+H⁺).

¹H NMR (CDCl₃): δ 8.33-8.30 (1H, m), 7.19-7.14 (1H, m), 7.04-7.03 (1H,m), 4.73-4.57 (1H, m), 3.82-3.71 (4H, m), 3.31 (2H, t, J=6.4 Hz),2.95-2.83 (2H, m), 2.41-2.29 (4H, m), 1.79-1.68 (2H, m), 1.67-1.25 (8H,m), 1.54 (6H, d, J=7.0 Hz).

Step 2.6-chloro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

The title compound was prepared according to the procedure described inStep 4 of Example 10 from6-chloro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide(Step 1 of Example 11)

MS (ESI) m/z: 465 (M+H⁺).

¹H NMR (DMSO-d₆): δ 8.84-8.76 (1H, m), 8.10-8.07 (1H, m), 7.51-7.45 (1H,m), 7.32-7.25 (1H, m), 5.38-5.32 (1H, m), 4.73-4.56 (1H, m), 3.70-3.55(6H, m), 3.41-2.91 (7H, m), 1.95-1.58 (8H, m), 1.48 (6H, d, J=7.7 Hz).

Anal. calcd. for C₂₃H₃₄N₄O₄Cl2.0.5H₂O: C, 54.12; H, 6.91; N, 10.98.Found: C, 53.85; H, 6.90; N, 10.78.

Example 125-chloro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

Step 1. 5-chloro-N-isopropyl-2-nitroaniline

The title compound was prepared according to the procedure described inStep 1 of Example 10 from 5-chloro-2-nitroaniline.

¹H NMR (CDCl₃): δ 8.12 (1H, d, J=9.2 Hz), 6.84 (1H, d, J=2.0 Hz), 6.57(1H, dd, J=9.2, 2.0 Hz), 3.81-3.71 (1H, m), 1.33 (6H, d, J=6.2 Hz)

Step 2. 6-chloro-1-isopropyl-1.3-dihydro-2H-benzimidazol-2-one

A mixture of 5-chloro-N-isopropyl-2-nitroaniline (Step 1 of Example 12,0.76 g, 3.54 mmol), iron (0.99 g, 17.7 mmol) and ammonium chloride (0.38g, 7.08 mmol) was suspended in ethanol (27 mL) and H₂O (9 mL). Then, themixture was heated at 80° C. for 3 h. After cooling, the insolublematerials was filtered off on a pad of Celite, and the filtrate wasevaporated under reduced pressure. To the residue was addedN,N′-carbonyldiimidazole (CDI, 0.57 g, 3.50 mmol) and THF (10 mL) andthen stirred at 100° C. for 10 h. After cooling, the volatile materialswere removed under reduced pressure and the residue was partitionedbetween ethylacetate and H₂O. After extraction with ethylacetate (3times), the combined organic phase was washed with brine, dried overMgSO₄ and concentrated. The residue was chromatographed on a column ofsilica gel eluting with hexane/ethyl acetate (2:1) to give 0.30 g (40%)of the title compound as a white solid.

¹H NMR (CDCl₃): δ 6.99-6.90 (2H, m), 6.84-6.74 (1H, m), 4.94-4.77 (1H,m), 1.64 (6H, d, J=7.0 Hz)

Step 3.5-chloro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

The title compound was prepared according to the procedure described inStep 3 of Example 9 from6-chloro-1-isopropyl-1,3-dihydro-2H-benzimidazol-2-one (Step 2 ofExample 12) and4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol (Step2 of Example 1).

MS (ESI) m/z: 465 (M+H⁺).

¹H NMR (CDCl₃): δ 8.88-8.78 (1H, m), 8.21-8.14 (1H, m), 7.19-7.10 (2H,m), 4.73-4.56 (1H, m), 3.87-3.69 (4H, m), 3.30 (2H, t, J=6.2 Hz),2.94-2.84 (2H, m), 2.41-2.27 (4H, m), 1.79-1.68 (2H, m), 1.67-1.25 (11H,m).

Anal. calcd. for C₂₃H₃₃N₄O₄Cl: C, 59.41; H, 7.15; N, 12.05. Found: C,59.27; H, 7.10; N, 11.72.

Example 13N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-5-methyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

Step 1. N-isopropyl-5-methyl-2-nitroaniline

The title compound was prepared according to the procedure described inStep 1 of Example 9 from 2-fluoro-4-methyl-1-nitrobenzene.

¹H NMR (CDCl₃): δ 8.12-8.01 (2H, m), 6.63 (1H, brs), 6.42 (1H, d, J=10.3Hz), 3.94-3.72 (1H, m), 2.33 (3H, s), 1.32 (6H, d, J=6.4 Hz)

Step 2. 1-isopropyl-6-methyl-1,3-dihydro-2H-benzimidazol-2-one

The title compound was prepared according to the procedure described inStep 2 of Example 9 from N-isopropyl-5-methyl-2-nitroaniline (Step 1 ofExample 13).

MS (ESI) m/z: 191 (M+H⁺).

¹H NMR (CDCl₃): δ 7.04-6.93 (2H, m), 6.90-6.80 (1H, m), 4.82-4.63 (1H,m), 2.40 (3H, s), 1.55 (6H, d, J=7.0 Hz).

Step 3.N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-5-methyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

The title compound was prepared according to the procedure described inStep 3 of Example 9 from1-isopropyl-6-methyl-1,3-dihydro-2H-benzimidazol-2-one (Step 2 ofExample 13) and4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol (Step2 of Example 1).

MS (ESI) m/z: 445 (M+H⁺).

¹H NMR (CDCl₃): δ 8.97-8.84 (1H, m), 8.10 (1H, d, J=8.8 Hz), 7.01-6.93(2H, m), 4.76-4.58 (1H, m), 3.85-3.69 (4H, m), 3.30 (2H, t, J=6.4 Hz),2.94-2.82 (2H, m), 2.41 (3H, s), 2.43-2.27 (4H, m), 1.80-1.68 (2H, m),1.67-1.25 (11H,

Anal. calcd. for C₂₄H₃₆N₄O₄: C, 64.84; H, 8.16; N, 12.60. Found: C,64.78; H, 8.29; N, 12.58.

Example 14N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-4-methyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

Step 1.N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-4-methyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

The title compound was prepared according to the procedure described inStep 3 of Example 9 from1-isopropyl-7-methyl-1,3-dihydro-2H-benzimidazol-2-one (I. Tapia et al.,J. Med. Chem., 42, 2880 (1999)) and4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol (Step2 of Example 1).

MS (ESI) m/z: 445 (M+H⁺).

¹H NMR (CDCl₃): δ 9.11-8.97 (1H, m), 8.17 (1H, d, J=7.7Hz), 7.10-6.88(2H, m), 4.99-4.82 (1H, m), 3.91-3.69 (4H, m), 3.29 (2H, t, J=6.2 Hz),2.94-2.82 (2H, m), 2.59 (3H, s), 2.43-2.27 (4H, m), 1.84-1.19 (7H, m),1.62 (6H, d, J=6.8 Hz).

Anal. calcd. for C₂₄H₃₆N₄O₄: C, 64.84; H, 8.16; N, 12.60. Found: C,64.73; H, 8.35; N, 12.56.

Example 15N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-4,5-dimethyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

Step 1. N-isopropyl-2,3-dimethyl-6-nitroaniline

The title compound was prepared according to the procedure described inStep 1 of Example 10 from 2,3-dimethyl-6-nitroaniline.

¹H NMR (CDCl₃): δ 7.82 (1H, d, J=8.6 Hz), 6.79 (1H, d, J=8.4 Hz),3.52-3.34 (1H, m), 2.30 (3H, s), 2.24 (3H, s), 1.11 (6H, d, J=6.2 Hz)

Step 2. 1-isopropyl-6,7-dimethyl-1,3-dihydro-2H-benzimidazol-2-one

The title compound was prepared according to the procedure described inStep 2 of Example 9 from N-isopropyl-2,3-dimethyl-6-nitroaniline (Step 1of Example 15).

¹H NMR (CDCl₃): δ 7.11 (1H, brs), 6.92-6.70 (1H, m), 5.00-4.82 (1H, m),2.45 (3H, s), 2.32 (3H, s), 1.63 (6H, d, J=7.0 Hz).

Step 3.N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-4,5-dimethyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

To a stirred mixture of1-isopropyl-6,7-dimethyl-1,3-dihydro-2H-benzimidazol-2-one (Step 2 ofExample 15, 204 mg, 1 mmol) and p-nitrophenylchloroformate (220 mg, 1.1mmol) in dichloromethane (7 mL) was added triethylamine (0.42 mL, 3.0mmol) at room temperature. After being stirred for 2 h, a solution of4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol (Step2 of Example 1, 230 mg, 1.0 mmol) in dichloromethane (3 mL) was added tothe mixture. After being stirred for 4 h, the mixture was diluted withethyl acetate (50 mL). Then, the organic layer was washed with 0.5 NNaOH aq. (5 mL) for 5 times and brine, dried over MgSO₄ andconcentrated. The residue was filtered through pad of aminopropyl-silicagel eluting with hexane/ethyl acetate (3:1) and the filtrate wasconcentrated. To the mixture was added 10% HCl-methanol (5 mL) wasstirred for 1 h. Then, the volatile components were removed underreduced pressure and the residue was recrystallized from ethanol-diethylether to give 100 mg (20%) of the title compound as a colorless powder.

MS (ESI) m/z: 459 (M+H⁺).

¹H NMR (DMSO-d₆): δ 8.96-8.87 (1H, m), 7.84 (1H, d, J=8.3 Hz), 6.95 (1H,d, J=8.3 Hz), 5.34-5.21 (1H, m), 5.01-4.86 (1H, m), 3.69-3.53 (6H, m),3.41-2.91 (7H, m), 2.45 (3H, s), 2.28 (3H, s), 1.87-1.70 (3H, m),1.67-1.48 (5H, m), 1.52 (6H, d, J=6.6 Hz).

Anal. calcd. for C₂₅H₃₉N₄O₄Cl.0.5H₂O: C, 59.57; H, 8.00; N, 11.12.Found: C, 59.53; H, 7.98; N, 11.10.

Example 166-fluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-5-methyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

Step 1. 4-fluoro-N-isopropyl-5-methyl-2-nitroaniline

The title compound was prepared according to the procedure described inStep 1 of Example 9 from 1,4-difluoro-2-methyl-5-nitrobenzene (T.Timothy et al., J. Med. Chem., 35, 2321 (1992)).

MS (ESI) m/z: 213 (M+H⁺).

¹H NMR (CDCl₃): δ 7.82 (1H, d, J=10.3 Hz), 6.64 (1H, d, J=6.4 Hz),3.88-3.67 (1H, m), 2.30 (3H, s), 1.31 (6H, d, J=6.4 Hz)

Step 2. 5-fluoro-1-isopropyl-6-methyl-1,3-dihydro-2H-benzimidazol-2-one

The title compound was prepared according to the procedure described inStep 2 of Example 9 from 4-fluoro-N-isopropyl-5-methyl-2-nitroaniline(Step 1 of Example 16).

MS (ESI) m/z: 209 (M+H⁺).

¹H NMR (CDCl₃): δ 7.00-6.96 (1H, m), 6.92-6.90 (1H, m), 4.75-4.56 (1H,m), 2.31 (3H, s), 1.55 (6H, d, J=7.0 Hz).

Step 3.6-fluoro-N-({1-{(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-5-methyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide

The title compound was prepared according to the procedure described inStep 3 of Example 9 from5-fluoro-1-isopropyl-6-methyl-1,3-dihydro-2H-benzimidazol-2-one (Step 2of Example 16) and4-{[4-(aminomethyl)piperidin-1-yl]methyl}tetrahydro-2H-pyran-4-ol (Step2 of Example 1).

MS (ESI) m/z: 463 (M+H⁺).

¹H NMR (CDCl₃): δ 8.92-8.83 (1H, m), 7.96 (1H, d, J=10.1 Hz), 6.91 (1H,d, J=6.2 Hz), 4.75-4.56 (1H, m), 3.85-3.70 (4H, m), 3.30 (2H, t, J=6.4Hz), 2.94-2.82 (2H, m), 2.42-2.29 (7H, m), 1.84-1.19 (7H, m), 1.55 (6H,d, J=7.0 Hz).

Step 4.6-fluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-5-methyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamidehydrochloride

The title compound was prepared according to the procedure described inStep 4 of Example 10 from6-fluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-5-methyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide(Step 3 of Example 16)

MS (ESI) m/z: 463 (M+H⁺).

¹H NMR (DMSO-d₆): δ 9.55-9.11 (1H, m), 8.89-8.74 (1H, m), 7.77 (1H, d,J=10.4 Hz), 7.40 (1H, d, J=6.6 Hz), 5.42-5.34 (1H, m), 4.70-4.56 (1H,m), 3.69-3.53 (6H, m), 3.52-2.91 (7H, m), 2.29 (3H, s), 1.87-1.70 (3H,m), 1.95-1.55 (8H, m), 1.48 (6H, d, J=6.8 Hz).

Anal. calcd. for C₂₄H₃₆N₄O₄FCl: C, 57.76; H, 7.27; N, 11.23. Found: C,57.47; H, 7.40; N, 11.05.

Preparation 1. Step 1. tert-Butyl4-({[(3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazol-1-yl)carbonyl]amino}methyl)piperidine-1-carboxylate

To a stirred solution of 1-isopropyl-1,3-dihydro-2H-benzimidazol-2one(J. Med. Chem. 1999, 42, 2870-2880) (3.00 g, 17.02 mmol) andtriethylamine (7.12 ml, 51.06 mmol) in 70 ml tetrahydrofuran was addedtriphosgen (5.15 g, 17.02 mmol) in 14 ml tetrahydrofuran at roomtemperature. The reaction mixture was refluxed for 19 hours. The mixturewas then cooled to room temperature, tert-butyl4-(aminomethyl)piperidine-1-carboxylate (J. Prugh, L. A. Birchenough andM. S. Egbertson, Synth. Commun., 1992, 22, 2357-60) (3.28 g, 15.32 mmol)in 10 ml tetrahydrofuran was added. The reaction mixture was refluxedfor another 24 hours. Then cooled and basified with aqueous saturatedNaHCO₃ 50 ml, and extracted with ethyl acetate 100 ml for three times.The combined extract was washed with brine, dried over MgSO₄ andconcentrated. Flash chromatography of the residue (elutent: hexane/ethylacetate=5/1 to 1/2) afforded a colorless oil 3.99 g (62%) as the titledcompound.

¹H-NMR (CDCl₃) δ: 9.04-8.88 (1H, m), 8.83-8.20 (1H, m), 7.26-7.10 (3H,m), 4.80-4.60 (1H, m), 4.28-4.02 (2H, m), 3.32 (2H, t, J=6.1 Hz),2.82-2.60 (2H, m), 1.94-1.10 (5H, m), 1.57 (6H, d, J=7.1 Hz), 1.45 (9H,s).

Step 2.3-Isopropyl-2-oxo-N-(piperidin-4-ylmethyl)-2,3-dihydro-1H-benzimidazole-1-carboxamide

A solution of tert-butyl4-({[(3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazol-1-yl)carbonyl]amino}methyl)piperidine-1-carboxylate(3.992 g, 9.58 mmol) in 50 ml 10% hydrochloric acid in methanol and 10ml concentrated hydrochloric acid was stirred at room temperature for 18hours. The mixture was then concentrated and basified with aqueousNa₂CO₃, extracted with CHCl₃ 100 ml for 3 times. The combined extractwas dried and concentrated. Flash chromatography of the residue(NH-silica gel, elutent: CH₂Cl₂/methanol=100/1) afforded a colorless oil2.272 g (75%) as the titled compound.

MS (ESI) m/z: 317 (M+H)⁺.

¹H-NMR (CDCl₃) δ: 8.93 (1H, br), 8.32-8.22 (1H, m), 7.24-7.02 (3H, m),4.80-4.61 (1H, m), 3.31 (2H, t, J=6.0 Hz), 3.20-3.05 (2H, m), 2.79-2.54(2H, m), 1.84-1.52 (3H, m), 1.57 (6H, d, J=6.9 Hz), 1.36-1.13 (2H, m).

Step 3.N-{[1-(3-Hydroxy-3-methyl-2-oxobutyl)piperidin-4-yl]methyl}-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide,monooxalate salt

A mixture of3-isopropyl-2-oxo-N-(piperidin-4-ylmethyl)-2,3-dihydro-1H-benzimidazole-1-carboxamide(250 mg, 0.790 mmol), 1-bromo-3-hydroxy-3-methylbutan-2-one (G. Bertram;A. Scherer; W. Steglich; W. Weber, Tetrahedron Lett., 1996, 37,7955-7958) (181 mg, 1.343 mmol) and triethylamine (0.28 ml, 1.975 mmol)in 8 ml tetrahydrofuran was refluxed for 15 hours. Then cooled anddiluted with 100 ml ethyl acetate and was washed with aqueous NaHCO₃ 20ml, brine, dried over MgSO₄ and concentrated. Flash chromatography ofthe residue (elutent: CH₂Cl₂/methanol=100/1 to 30/1) afforded acolorless oil 202 mg (61%). The oil (202 mg) was dissolved in 3 mlmethanol and acidified with a solution of 44 mg oxalic acid in 1 mlMeOH. The mixture was concentrated. Recrystallization of the resultingsolid with EtOH—AcOEt afforded a white solid 246 mg as the titledcompound.

MS (ESI) m/z: 417 (M+H)⁺.

m.p.: 140.5° C.

IR (KBr) v: 3404, 3306, 2980, 2941, 1728, 1690, 1612, 1541 cm⁻¹.

¹H-NMR (CDCl₃) (free base) δ: 8.90 (1H, br) 8.30-8.20 (1H, m), 7.24-7.10(3H, m), 4.78-4.61 (1H, m), 3.37 (2H, s), 3.33 (2H, t, J=6.3 Hz),3.00-2.86 (2H, m), 2.22-2.06 (2H, m), 1.90-1.22 (5H, m), 1.57 (6H, d,J=7.0 Hz), 1.35 (6H, s).

¹H-NMR (DMSO-d₆) (salt form) δ: 8.92-8.81 (1H, m) 8.07 (1H, dd, J=7.7,6.8 Hz), 7.44 (1H, d, J=7.7 Hz), 7.28-7.10 (2H, m), 4.74-4.60 (1H, m),4.36 (2H, bv), 4.00-2.70 (6H, m), 1.90-1.44 (5H, m), 1.49 (6H, d, J=6.4Hz), 1.24 (6H, s).

Anal. Calcd. for C₂₄H₃₄N₄O₈.0.3C₂H₆O.1H₂O: C, 54.88; H, 7.08; N, 10.41.Found: C, 55.26; H, 7.18; N, 10.07.

All publications, including but not limited to, issued patents, patentapplications, and journal articles, cited in this application are eachherein incorporated by reference in their entirety.

Although the invention has been described above with reference to thedisclosed embodiments, those skilled in the art will readily appreciatethat the specific experiments detailed are only illustrative of theinvention. It should be understood that various modifications can bemade without departing from the spirit of the invention. Accordingly,the invention is limited only by the following claims

1. A compound of the formula (I):

wherein Het represents a heterocyclic group having one nitrogen atom, towhich B binds directly, and from 4 to 7 carbon atoms, and saidheterocyclic group being unsubstituted or substituted by 1 to 4substituents independently selected from the group consisting ofsubstituents α¹; A represents an alkylene group having from 1 to 4carbon atoms; B represents a covalent bond or an alkylene group havingfrom 1 to 5 carbon atoms, and said alkylene group being unsubstituted orsubstituted by an oxo group when R³ represents a heterocyclic group; R¹represents an isopropyl group or a cyclopentyl group; R² independentlyrepresents a halogen atom or an alkyl group having from 1 to 4 carbonatoms; m is 0, 1, 2, 3 or 4; and R³ represents (i) a cycloalkyl grouphaving from 3 to 8 carbon atoms, and said cycloalkyl group beingsubstituted by 1 to 5 substituents independently selected from the groupconsisting of substituents α², or (ii) a heterocyclic group having from3 to 8 atoms, and said heterocyclic group being unsubstituted orsubstituted by 1 to 5 substituents independently selected from the groupconsisting of substituents β, said substituents α¹ are independentlyselected from a hydroxy group and an amino group; said substituents α²are independently selected from a hydroxy group, an amino group, ahydroxy-substituted alkyl group having from 1 to 4 carbon atoms, acarboxyl group and an alkoxy group having from 1 to 4 carbon atoms; andsaid substituents β are independently selected from a hydroxy group, ahydroxy-substituted alkyl group having from 1 to 4 carbon atoms, acarboxyl group, an amino group, an alkyl group having from 1 to 4 carbonatoms, an amino-substituted alkyl group having from 1 to 4 carbon atomsand a carbamoyl group, or a pharmaceutically acceptable salts thereof.2. The compound or the pharmaceutically acceptable salt thereof, asclaimed in claim 1, wherein Het represents a heterocyclic group selectedfrom

said heterocyclic group being unsubstituted or substituted by 1 to 3substituents independently selected from the group consisting ofsubstituents α¹; and A represents an alkylene group having from 1 to 3carbon atoms.
 3. The compound or the pharmaceutically acceptable saltthereof, as claimed in claim 1, wherein Het represents a group offormula

and this group being unsubstituted or substituted by one substituentselected from the group consisting of substituents α¹; A represents analkylene group having from 1 to 2 carbon atoms; B represents an alkylenegroup having from 1 to 4 carbon atoms, and said alkylene group beingunsubstituted or substituted by an oxo group when R³ represents aheterocyclic group; R² independently represents a halogen atom or analkyl group having from 1 to 2 carbon atoms; m is 0, 1 or 2; and R³represents (i) a cycloalkyl group having from 4 to 7 carbon atoms, andsaid cycloalkyl group being substituted by 1 to 3 substituentsindependently selected from the group consisting of substituents α², or(ii) a heterocyclic group having from 4 to 7 atoms, and saidheterocyclic group being unsubstituted or substituted by 1 to 3substituents independently selected from the group consisting ofsubstituents β.
 4. The compound or the pharmaceutically acceptable saltthereof, as claimed in claim 1, wherein Het represents a group offormula

and this group being unsubstituted or substituted by one substituentselected from the group consisting of substituents α¹; A represents amethylene group; B represents an alkylene group having from 1 to 2carbon atoms; R¹ represents an isopropyl group; R² independentlyrepresents a fluorine atom, a chlorine atom or a methyl; and R³represents (i) a cycloalkyl group having from 5 to 7 carbon atoms, andsaid cycloalkyl group being substituted by 1 to 2 substituentsindependently selected from the group consisting of substituents α², or(ii) a heterocyclic group having from 5 to 7 atoms, and saidheterocyclic group being unsubstituted or substituted by 1 to 2substituents independently selected from the group consisting ofsubstituents β, said substituents α² are independently selected from ahydroxy group, an amino group and an alkoxy group having from 1 to 2carbon atoms; and said substituents β are independently selected from ahydroxy group, a hydroxy-substituted alkyl group having from 1 to 2carbon atoms, a carboxyl group, an amino group, an amino-substitutedalkyl group having from 1 to 2 carbon atoms and a carbamoyl group. 5.The compound or the pharmaceutically acceptable salt thereof, as claimedin claim 1, wherein Het represents a group of formula

A represents a methylene group; B represents a methylene group; R¹represents an isopropyl group; R² represents a fluorine atom; m is 0 or1; and R³ represents (i) a cycloalkyl group having from 5 to 6 carbonatoms, and said cycloalkyl group being substituted by 1 to 2substituents independently selected from the group consisting ofsubstituents α², or (ii) a heterocyclic group having from 5 to 6 atoms,and said heterocyclic group being unsubstituted or substituted by 1 to 2substituents independently selected from the group consisting ofsubstituents β, said substituents α² are independently selected from ahydroxy group and an amino group; and said substituents β areindependently selected from a hydroxy group and an amino group.
 6. Thecompound or the pharmaceutically acceptable salt thereof, as claimed inclaim 1, wherein Het represents a group of formula

A represents a methylene group; B represents a methylene group; R¹represents an isopropyl group; R² represents a fluorine atom; m is 0;and R³ represents (i) a cyclohexyl group substituted by 1 to 2substituents independently selected from a hydroxy group or an aminogroup, or (ii) a heterocyclic group having from 6 atoms, and saidheterocyclic group being substituted by a hydroxy group or an aminogroup.
 7. The compound or the pharmaceutically acceptable salt, asclaimed in claim 6, wherein R³ represents (i) a cyclohexyl groupsubstituted by 1 or 2 hydroxy group, or (ii) a tetrahydropyran groupsubstituted by 1 or 2 hydroxy group.
 8. The compound or thepharmaceutically acceptable salt, as claimed in claim 7, wherein R³represents hydroxytetrahydropyranyl or dihydroxycyclohexyl.
 9. Thecompound as claimed in claim 1, which is selected from:N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide;N-({1-[(trans-1,4-dihydroxyhexyl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide;N-({1-[(cis-1,4-dihydroxyhexyl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide;and6-fluoro-N-({1-[(4-hydroxytetrahydro-2H-pyran-4-yl)methyl]piperidin-4-yl}methyl)-3-isopropyl-2-oxo-2,3-dihydro-1H-benzimidazole-1-carboxamide,or the pharmaceutically acceptable salt thereof.
 10. The compound or thepharmaceutically acceptable salt, as claimed in any one of claims 1 to9, wherein the pharmaceutically acceptable salt is a hydrochloride orhemiedysilate.
 11. A pharmaceutical composition including the compoundor the pharmaceutically acceptable salt thereof, as claimed in any oneof claims 1 to 9, together with a pharmaceutically acceptable excipient.12. A method of the treatment of disease conditions mediated by 5-HT₄receptor activity, in a mammalian subject, which comprises administeringto the said subject in need of such treatment a therapeuticallyeffective amount of the compound or the pharmaceutically acceptable saltthereof, as claimed in any one of claims 1 to
 9. 13. A method of thetreatment of diseases selected from gastroesophageal reflux disease,gastrointestinal disease, gastric motility disorder, non-ulcerdyspepsia, functional dyspepsia, irritable bowel syndrome (IBS),constipation, dyspepsia, esophagitis, gastroesophageral disease, nausea,central nervous system disease, Alzheimer's disease, cognitive disorder,emesis, migraine, neurological disease, pain, cardiovascular disorders,cardiac failure, heart arrhythmia, diabetes and apnea syndrome, whichcomprises administering to the said subject a therapeutically effectiveamount of the compound or the pharmaceutically acceptable salt thereof,as claimed in any one of claims 1 to
 9. 14. Use of the compound or thepharmaceutically acceptable salt thereof, as claimed in any one ofclaims 1 to 9 for the manufacture of a medicament for the treatment ofdisease conditions mediated by 5-HT₄ receptor activity, in a mammaliansubject.
 15. Use of a compound as claimed in claim 14, wherein saidcondition is selected from gastroesophageal reflux disease,gastrointestinal disease, gastric motility disorder, non-ulcerdyspepsia, functional dyspepsia, irritable bowel syndrome (IBS),constipation, dyspepsia, esophagitis, gastroesophageral disease, nausea,central nervous system disease, Alzheimer's disease, cognitive disorder,emesis, migraine, neurological disease, pain, cardiovascular disorders,cardiac failure, heart arrhythmia, diabetes and apnea syndrome.
 16. Acompound of the formula (2-A′):

wherein R^(a) represents a hydrogen atom or a N-protecting group; Hetrepresents a heterocyclic group having one nitrogen atom, to which Bbinds directly, and from 4 to 7 carbon atoms, and said heterocyclicgroup being unsubstituted or substituted by 1 to 4 substituentsindependently selected from the group consisting of substituents α¹; Arepresents an alkylene group having from 1 to 4 carbon atoms; Brepresents a covalent bond or an alkylene group having from 1 to 5carbon atoms, and said alkylene group being unsubstituted or substitutedby an oxo group when R³ represents a heterocyclic group; R³ represents(i) a cycloalkyl group having from 3 to 8 carbon atoms, and saidcycloalkyl group being substituted by 1 to 5 substituents independentlyselected from the group consisting of substituents α², or (ii) aheterocyclic group having from 3 to 8 atoms, and said heterocyclic groupbeing unsubstituted or substituted by 1 to 5 substituents independentlyselected from the group consisting of substituents β, said substituentsα¹ are independently selected from a hydroxy group and an amino group;said substituents α² are independently selected from a hydroxy group, anamino group, a hydroxy-substituted alkyl group having from 1 to 4 carbonatoms, a carboxyl group and an alkoxy group having from 1 to 4 carbonatoms; and said substituents β are independently selected from a hydroxygroup, a hydroxy-substituted alkyl group having from 1 to 4 carbonatoms, a carboxyl group, an amino group, an alkyl group having from 1 to4 carbon atoms, an amino-substituted alkyl group having from 1 to 4carbon atoms and a carbamoyl group, or a salt thereof
 17. The compoundor the salt thereof, as claimed in claim 16, wherein R^(a) represents ahydrogen atom or a t-butoxycarbonyl group; Het represents a group offormula

A represents an methylene group; B represents an methylene group; and R³represents hydroxytetrahydropyranyl or dihydroxycyclohexyl.
 18. Acombination of the compound or the pharmaceutically acceptable saltthereof as claimed in any one of claims 1 to 9, and anotherpharmacologically active agent.
 19. A pharmaceutical compositionincluding the compound or the pharmaceutically acceptable salt thereofas claimed in any one of claims 1 to 9, and another pharmacologicallyactive agent.
 20. A compound of the formula (I):

wherein Het represents a heterocyclic group having one nitrogen atom, towhich B binds directly, and from 4 to 7 carbon atoms, and saidheterocyclic group being unsubstituted or substituted by 1 to 4substituents independently selected from the group consisting ofsubstituents α¹; A represents an alkylene group having from 1 to 4carbon atoms; B represents a covalent bond or an alkylene group havingfrom 1 to 5 carbon atoms, and said alkylene group being unsubstituted orsubstituted by an oxo group when R³ represents a heterocyclic group; R¹represents an isopropyl group or a cyclopentyl group; R² independentlyrepresents a halogen atom or an alkyl group having from 1 to 4 carbonatoms; m is 0, 1, 2, 3 or 4; and R³ represents (i) a cycloalkyl grouphaving from 3 to 8 carbon atoms, and said cycloalkyl group beingsubstituted by 1 to 5 substituents independently selected from the groupconsisting of substituents α², or (ii) a heterocyclic group having from3 to 8 atoms, and said heterocyclic group being unsubstituted orsubstituted by 1 to 5 substituents independently selected from the groupconsisting of substituents β, said substituents α¹ are independentlyselected from a hydroxy group and an amino group; said substituents α²are independently selected from a hydroxy group, an amino group, ahydroxy-substituted alkyl group having from 1 to 4 carbon atoms, acarboxyl-substituted alkyl group having 1 to 4 carbon atoms, a carboxylgroup and an alkoxy group having from 1 to 4 carbon atoms; and saidsubstituents β are independently selected from a hydroxy group, ahydroxy-substituted alkyl group having from 1 to 4 carbon atoms, acarboxyl-substituted alkyl group having 1 to 4 carbon atoms, a carboxylgroup, an amino group, an alkyl group having from 1 to 4 carbon atoms,an amino-substituted alkyl group having from 1 to 4 carbon atoms and acarbamoyl group, or a pharmaceutically acceptable salts thereof.